CZ2000632A3 - Derivatives of 4-cyano-4 deformylsordarin, process of their preparation and pharmaceutical and agrochemical preparations in which these derivative are comprised - Google Patents

Abstract

The 4-cyano-4-deformylsordarin derivatives are fungicidal agents suitable for the treatment and / or prevention of human and animal fungal infections and for controlling phytopathogenic fungi agricultural crops.

Description

Derivative ^. 4-cyano-4-deformylsordarin ₍ / and an agrochemical composition comprising the following methods of manufacture | pharmaceuticals and derivatives

Field of technology

The present invention relates to 4-cyano-4-deformylsordarin derivatives which are potent fungicidal agents with a broad spectrum of action and enhanced stability, to processes for the preparation of these derivatives, to pharmaceutical and agricultural compositions containing these compounds and to controlling fungal infections in humans, animals and plant materials. of these compounds.

Prior art

Sordarin is a fungicidal antibiotic isolated from the fungus Sordaria araneosa (see British Patent GB 1162027 and Helvetica Chimica Acta, 1971, 51, 119-120). Other fungicidal agents having the same structural basis as sordarin are also known. Kokai J62040292 describes a compound called zofimarin which was isolated from Zofiela marina sp .; Kokai J06157582 discloses a compound designated BE-31405 which was | isolated from Penicillium sp .; and in J. Antibiotics, 1995, 48,

1171-1172 discloses a compound designated SCH57404. Ve &

PCT published applications WO96 / 14326 and WO96 / 14327 disclose semisynthetic sordarin derivatives.

The essence of the invention

The present invention relates to compounds of formula (I):

(AND)

wherein Z is a tetrahydropyran group selected from the group consisting of compounds of formulas (a) to (f):

(a) (b) (c)

í, t

and their salts and solvates (eg hydrates) or their metabolically labile derivatives, wherein

R ^a is C (O) CH ₃ or methyl;

R ¹ is a hydrogen atom, a halogen atom, a hydroxyl group, an alkoxy group having 1 to 4 carbon atoms or an acyloxy group;

R ² and R ³ are each independently a hydrogen atom, an alkyl group having 1 to 6 carbon atoms or an alkyl group • · · · • ·

• · • · • · • · v

containing 1 to 4 carbon atoms substituted by an alkoxy group containing 1 to. 4 carbon atoms, or

R ² and R ³ together with the carbon atom to which they are attached are a carbonyl group (C = O), a thiocarbonyl group (C = S) or a cycloalkyl group containing 3 to 8 carbon atoms;

R ⁴ is a hydrogen atom or a CH ² R ⁷ group (wherein R ⁷ is a hydrogen atom, a hydroxyl group, an alkoxy group having 1 to 4 carbon atoms or an OCOR ⁸ group in which R ⁸ is an alkyl group having 1 to 4 carbon atoms or an aryl group);

R ⁵ and R ⁶ are each independently a hydrogen atom, an alkyl group having 1 to 6 carbon atoms or an alkoxy group having 1 to 4 carbon atoms substituted by an alkyl group having 1 to 4 carbon atoms, or

R ⁵ and R ⁶ together with the carbon atom to which they are attached are a carbonyl group (C = O), a thiocarbonyl group (C = S) or a cycloalkyl group containing 3 to 8 carbon atoms;

n is zero or 1;

X and Y are each independently oxygen, sulfur or CR ⁹ R ¹⁰ (wherein R ⁹ and R ¹⁰ are each independently hydrogen, alkyl of 1 to 6 hydrogen atoms, alkoxy of 1 to 4 hydrogen atoms or alkyl of 1 to 4 carbon atoms substituted by an alkoxy group having 1 to 4 carbon atoms, or R ⁹ and R ¹⁰ together with the carbon atom to which they are attached are a carbonyl group (C = O), a thiocarbonyl group (C = S), a cycloalkyl group containing 3 up to 8 carbon atoms or a group C = CHR, where R is a hydrogen atom or an alkyl group containing 1 to 4 carbon atoms); or when X or Y is an oxygen atom and ⁿ = 0, the groups -Y-CR ² R ³ and -X-CR ² R ^{3 respectively} may also be a group -N = CR ³ or a group -NR ¹² -CR ² R ³ - (where CR ² and R ³ are a carbonyl group (C = O) and R ¹² is an alkyl group containing 1 to 4 carbon atoms, • · ♦ · • »· ·

I í * »•» Mí • · * • «· ·« · * * »* * * * í» »* * • · · ·» * »•» »· · β · · * • # * * * * »» * • t · · k-

the acyl group COR ¹³ , where R ¹³ is an alkyl group containing 1 to 6 carbon atoms) or when Y is an oxygen atom and n = 0, X may be a group CR ¹¹ (where R ¹¹ is as defined above) which is bonded by a double bond to a pyran ring;

R ¹⁵ is a hydrogen atom, a halogen atom, an azido group, an alkyl group having 1 to 6 carbon atoms, a hydroxy group, an alkoxy group having 1 to 6 carbon atoms (optionally substituted by 1 or 2 hydroxy groups or their ketal or 1 or 2 alkoxy groups having 1 to 3 carbon atoms) ), an aryl-substituted alkoxy group having 1 to 4 carbon atoms, an alkenyloxy group having up to 6 carbon atoms, an OCOR ¹⁸ group (wherein R ¹⁸ is an aryl-substituted alkoxy group having 1 to 4 carbon atoms or an alkyl group having 1 to 10 carbon atoms optionally containing one or two double bonds) or an alkoxy group having 1 to 6 carbon atoms substituted by an alkoxy group having 1 to 4 carbon atoms and R ¹⁶ is a hydrogen atom or R ¹⁵ R ¹⁶ together with the carbon atom to which they are attached are a carbonyl group (C = O) or C = CH ₂ ;

R ¹⁷ is CH ₂ R ¹⁹ , wherein R ¹⁹ is hydrogen, hydroxyl, alkoxy having 1 to 14 carbon atoms, or OCOR ²⁰ , wherein R ²⁰ is alkyl having 1 to carbon atoms; and

W is an oxygen atom, a sulfur atom or a CH ₂ group; and the dotted line in group (a) indicates the possible presence of another bond;

R ^1a is a hydrogen atom, a halogen atom, a hydroxyl group or an alkoxy group having 1 to 4 carbon atoms

R ^2a is a hydrogen atom, a halogen atom, a hydroxyl group or an alkoxy group having 1 to 10 carbon atoms, an alkylthio group having 1 to 10 carbon atoms, an alkoxy group

• • · 1 to 6 carbon atoms substituted alkoxy group having 1 to 4 carbon atoms, aryl substituted alkoxy group having 1 to 6 carbon atoms, aryl substituted alkenyloxy group having 3 to 6 carbon atoms, azido group, NR ^5a group COR ^5a (wherein each R ^5a is independently hydrogen or alkyl of 1 to 6 carbon atoms), OR ^5a (wherein R ^6a is a cyclic ether having 4 to 8 atoms attached to the oxygen atom by a carbon atom of that cyclic ether adjacent to the oxygen atom ) or a group Y ^and C (= O) -X ^and -R ^7a , wherein Y ^a is an oxygen atom, a sulfur atom or an NH group, X ^a is either a bond, an oxygen atom or an NR ⁸ group in which the substituent R ^8a is an atom hydrogen or an alkyl group containing 1 to 6 carbon atoms and R ^7a is an alkyl group containing 1 to 10 carbon atoms optionally containing one or two double bonds, an aryl group, an aryl-substituted alkyl group containing having 1 to 4 carbon atoms, an aryl-substituted alkenyl group having 2 to 4 carbon atoms, a haloalkyl group having 1 to 6 carbon atoms or an alkoxy group having 1 to 6 carbon atoms substituted an alkyl group having 1 to 4 carbon atoms and R ^3a is a hydrogen atom or _R 2a _{and R} 3a _{S of} the carbon atom to which they are attached are a carbonyl group (C = O) or a group C = NOR ^9a (wherein R ^9a is an alkyl group having 1 to 6 carbon atoms); and

R ^4a is a hydroxyl group, an alkoxy group having 1 to 6 carbon atoms or an OC (= O) R ^{7a group} (wherein R ^7a is as defined above).

The present invention also includes a compound of formula (I) wherein Z is • · • · • · • ·

Another aspect of the invention is a pharmaceutical composition which comprises a fungicidally effective amount of a compound of formula (I) and a pharmaceutically acceptable carrier. The invention also relates to a pharmaceutical composition which is prepared by combining a compound of formula (I) and a pharmaceutically acceptable carrier.

Another aspect of the invention is also an agricultural composition comprising a fungicidally effective amount of a compound of formula (I) and an agriculturally acceptable carrier. The invention also relates to an agricultural composition which is prepared by combining a compound of formula (I) and an agriculturally acceptable carrier.

The present invention further relates to a method of treating a fungal infection in an animal (including a human) which comprises administering to a animal in need of such treatment a fungicidally effective amount of a compound of formula (I).

Another aspect of the present invention is a method of controlling phytopathogenic fungi of plants, which comprises applying a fungicidally effective amount of a compound of formula (I) to said plants.

Unless otherwise indicated, the terms used in this description of the invention have the following meanings:

The term "control or" control includes prophylactic use (ie, protection against infection) and therapeutic use (ie, eradication of infection).

The term "plants" includes all or part of the live plants, leaves, flowers, seeds, fruits and other plant material. Due to the possibility of applying the active ingredient to the soil, the term also includes the roots of the plant.

The term 'product' means an agricultural or agrochemical product that includes a product comprising active · ♦ • · 0 · • · · · • 0 000 • · «

00 • 0 * ·· 0

component (s), an inert carrier-forming component (s), and any product formed, directly or indirectly, by combination, complexation, or grouping of any two or more components or resulting from the dispersion of one or more components or other reactions or interactions of one or more components. Accordingly, a composition of the present invention includes any composition made by mixing a compound of the present invention and an agriculturally acceptable carrier.

The term "alkyl group" used alone as part of a group designation includes a linear or branched alkyl chain such as methyl group, ethyl group, n-propyl group, n-butyl group, isopropyl group, sec-butyl group, tert-butyl group, n- hexyl group and n-octyl group.

The term "aryl" used alone as part of the designation includes a phenyl group or a heteroaryl group, both of which may be optionally substituted with one to three groups independently selected from the group consisting of halogen, hydroxyl, alkyl of 1 to 6 carbon atoms, alkoxy containing 1 to 6 carbon atoms or an alkoxycarbonyl group containing 1 to 4 carbon atoms in the side chain. The heteroaromatic group may be a five- or six-membered heteroaromatic ring containing one or more heteroatoms selected from the group consisting of a nitrogen atom, an oxygen atom and a sulfur atom. Examples of suitable heteroaromatic groups include pyridyl, furyl, thienyl and pyrrolyl.

"Halogen atom or" halogen means a fluorine atom, a chlorine atom, a bromine atom or an iodine atom.

9 «♦ 9 9

9

999 9 <

When R ^{1 is} an acyloxy group, it may be, for example, OCOR ¹³ , in which R ^{13 is} as defined above.

A group of suitable salts of a compound of formula (I) includes basic inorganic salts such as alkali metal salts (e.g. sodium or potassium salts), ammonium salts and organic base salts. A group of suitable basic organic salts includes salts of amines such as trialkylamine (e.g. triethylamine), or salts of dialkylamines (e.g. dicyclohexylamine), salts of optionally substituted benzylamine (e.g. phenylbenzylamine or p-bromobenzylamine), salts of ethanolamine, diethanolamine, N-methylglucosamine, N- methylpiperidine, pyridine and substituted pyridine (e.g. collidine, lutidine, 4-methylaminopyridine), tri (hydroxymethyl) methylamine salts and amino acid salts (e.g. lysine or arganine).

Metabolically labile derivatives of a compound of formula (I) are compounds which are converted into a compound of formula (I) in a treated object (which may be an animal, plant (including leaves, flowers, fruits, seeds and other parts or fruits of a plant) or soil) ). Examples of such derivatives include commonly metabolically labile esters formed by carboxyl groups in the molecule.

Preparation of compounds

Compounds of formula (I) may be prepared from sordarin and its derivatives, sordaricin and other compounds of the sordarin type, all of which are described in the literature.

Sordarin is the acid [IR- (1a, 3aE, 4β, 4a3, 7β, 7aa, 8ap)] -8a [(6-deoxy-4-O-methyl-3-D-altropyranosyloxy) methyl] -4-formyl4, 4a, 5,6,7,7a, 8,8a-octahydro-7-methyl-3- (1-methylethyl) -1,4-methano-s-indacene-3a (1H) -carboxylic formula (II):

• ftftftft ft * ···· »· · ft ft · · · • ft ftft» ftft <

► ftft «» · · <

»· ·«

MeO

(II)

Sordarin can be obtained by culturing Sordaria araneosa NRRL 3196 (also deposited with the ATCC under the designation ATCC 36386) as described in British Patent GB 1162027 or in published PCT application WO96 / 14326. Sordarin can also be obtained by fermentation of Rosellinia subiculata and an unidentified fungus designated ATCC 74387 as described below.

Zofimarin can be obtained by fermentation of Zofiela marina SANK 21274 broth (ATCC 34456) as described in Kokai 62040292. As described in Kokai 06157582, a compound of formula I is wherein A is (f) and R ^a is an acetyl group, designated BE31405 as a product Penicillium sp. F31405. As reported in J. Antibiotics, 1995, 48 (10), 1171-1172, is a compound of formula (I) wherein A is (f) and R ^a is a methyl group, designated SCH57404 as a product of the fungus referred to in the Schering Culture Collection designation SCF1082A.

PCT Published Application WO96 / 14326 discloses a starting material for the preparation of sordarin derivatives of formula (I) wherein Z is (a) or (b) and PCT Published Application WO96 / 14327 discloses a starting material for the preparation of sordarin derivatives of formula (I). where Z is (c).

Sordaricin is an acid [IR- (Ια, 3ββ, 4β, 4ap, 7β, 7aa, 8ap)] - 4-formyl-8a- (hydroxymethyl) -4,4a, 5,6,7,7a, 8,8a-octahydro-7methyl -3- (1-methylethyl) -1,4-methano-s-indacene3a (1H) carboxylic formula (VI):

• 0 00 »· 0 0 ··: ::. · ··· »0 000 0 ·· · · 0« 00 *

(VI)

Sordaricin can be prepared from sordarin by reaction with concentrated hydrochloric acid. As described in published PCT application WO96 / 14326, sordaricin can also be obtained by fermentation of a mutant derived from Sordaria araneosa NRRL 3196 and biotransformation of sordarin using Koryneforms.

As mentioned above, two other sordarin-producing organisms were found.

One of the strains of sordarin-producing fungi is the unidentified sterile fungus GB3109, which was isolated from the internal tissues of the roots of the mangrove shrub Conocarpus erectus (Combretacea), which is stored in the collection of Manglar de Río Rincón, Península de Osa, Provincia de Puntarenas, as MF6232 in the culture collection of Merck & Co., Inc., Rahway, New Jersey. This culture was deposited on August 27, 1996 at the American Type Culture Collection, 12301 Parklawn Drive,

Maryland 20852, USA under the terms of the Budapest International Recognition of the Deposit of Microorganisms patent pending and designated ATCC 74387.

This mold was cultured on different mycological media under different light regimes and on leaves and filter paper, but in all of them it was not possible to obtain reproductive structures, so that it was not possible to identify this mold.

Rockville, contracts for the purpose of sterilized cases with • fe fefefefe fe «• · · • fefe • · • · • fe • fe«

fefefe «• · · fe fefefefe fe · • · fefe fefefe • fe ·· • fefe · fe fefe« fe fefe · • · · * fefe ··

Colonies of this fungus had the following morphology when grown on agar:

Colonies on oat agar (Difco) at 23 ° C and a 12 hour photoperiod grew slightly fast and reached a size of 85-90 millimeters after 14 days. The colonies had a sessile zone of progression, were flat, dark striped, strongly radially furrowed, with sessile moist mycelium in the center, silky at the edges with protruding creeping hyphal bundles or fibers. The colonies were translucent to pinkish (close to the shades referred to by Ridgway, R: Color Standards and Nomenclature, Washington, DC, 1912 as Pale Ochraceous Salmon, Light Ochraceous Salmon), pink-gray (referred to as Avellaneous, Cinnamon-Drab), or white in areas of the uppermost aerial mycelium. The back of the colonies was pink-gray. No exudates were observed and the smell of the colonies was neutral. No growth was observed on oat agar at 37 ° C.

Colonies on agar prepared from V8 juice (see Stevens, RB: Mycology Guidebook, University of Washington Press, Seattle, 1981, p. 665) grew slowly at 23 ° C and a 12-hour photoperiod and reached sizes 37-42 after 14 days. millimeters. The edges of the colonies were submerged and the mycelium was mostly sessile with a limited flaky aerial mycelium towards the outer third. The colonies were striped, translucent to slightly gray-pink, colored similar to oat agar culture, the reverse of the colonies was transparent to slightly reddish brown (close to the shades referred to in the above-mentioned publication as Wood Brown, Fawn Color).

Colonies on corn agar (Difco) at 25 ° C and a 12 hour photoperiod grew slowly and after 14 days reached • 9 ··· ♦ • 9

9 * 9

9 «

99 «9 9

9 9 9

9 · 9 • 9 9 9 • 9 99 sizes 33-34 millimeters. The edges of the colonies were submerged and the colonies from which air hyphae were released were striped and translucent.

The mycelium was formed by a highly branched, simply septal strain of hyaline hyphae.

The second sordarin-producing strain of fungi (designated GB3719) is Rosellinia subiculata (Ascomycotina, Xylariaceae) designated MF6239 in the Merck & Co., Inc., Rahway, New Jersey culture collection. This culture was deposited on 27 August 1996 at the Permanent American Type Culture Collection, 12301 Parklawn Drive, Rockville, Maryland 20852, USA under the terms of the Budapest Treaty on the International Recognition of the Deposit of Microorganisms for the Purposes of Patent Procedure and designated ATCC 74386.

Rosellinia subiculata ascombates (GB3719) were found on debarked hardwood branches on the banks of the Navesink River, Monmouth Co. in New Jersey. Using a sterilized microtome blade, all pieces of several ascomats, sacs, paraphyses, and ascospores were removed from their centers in the laboratory, removed with an insect attachment pin, and inoculated with malt fermentation extract agar. The ascospores were incubated overnight until germinated and transferred to malt fermentation agar agar to establish pure colonies.

The morphology of Rosellinia subiculata (GB3719) has generally confirmed the literature descriptions (Ellis, J. B. and Everhart, A. M. The North American Pyrenomycetes, published by Newfield, New Jersey, 1892, pp. 165-166; Petřini, L.

E. Rosellinia species of- the temperate zone. Sydowia, 1991, 44, 169-281). The key features that led to the determination of Rosellinia subiculata were: stromatic ascomatics occurring sporadically but clustered or sintered into small clusters on ···· ·· '• ··· * * · • · · · · · · 4 ·· ··· ·· ¹ · * ·· • »4 *» • · · · · • · ··· · · «· · • 4 ·· mycelial bed on the hemispherical, papillary, the mycelium was black; thin debarked wood; the fine, glossy, mycelial bedrock was dull, yellowish, or sometimes appeared only as light-colored discolored wood adjacent to the fungus; the sacs were cylindrical with a starchy apical stopper; the ascospores were brown-gray, broadly elliptical to slightly kidney-shaped, fine without pendants or coatings, with a straight, lower cleft slit, and measured 10-12 x 5-6 micrometers.

Colonies of this fungus had the following morphology when grown on agar:

Colonies on oat agar at 23 ° C and a 12-hour photoperiod grew slightly rapidly and reached 73-75 millimeters in size after 14 days. The colonies had a sessile zone of progression, were flat, dark striped, the inner third of the mycelia was white velvety to flaky, the outer two thirds of the mycelia were moist and sessile. The colonies were translucent to white or pinkish, slightly burgundy pink on the reverse (close to the shade referred to in the above-mentioned publication as Light Vinaceous Cinnamon). No exudates were observed and the colonies smelled faintly. No growth was observed on oat agar at 37 ° C.

Colonies on agar prepared from V8 juice at 23 ° C and a 12-hour photoperiod grew slowly and reached a size of 25-35 millimeters after 14 days. The edges of the colonies were submerged and the mycelium was mostly flaky aerial mycelium towards the Colonies were striped, translucent (Vinaceous Cinnamon), the reverse of the colonies was transparent to cinnamon (close to the shades described in the above publication as sessile with limited to the inner third, to slightly gray-pink

44 * · · · • · · • • 444

4444

4 4

4 44 * · · *

4 · · · »*» 4 44

4 4

4 4

4 4

4 4

44

Orange-Cinnamon, Cinnamon), or faintly reddish-brown (Russet, Fawn Color), smelly.

Colonies on corn agar at 25 ° C and a 12 hour photoperiod grew slowly and reached a size of 29-34 millimeters after 14 days. The edges of the colonies were submerged and from the air hyphae-releasing colony they were striped and translucent or had a reduced white mycelium at the point of inoculation, the reverse of the colonies was colorless.

When first grown in culture in August 1993, the strain produced only a limited number of conidiophores and Geniculosporium conidia anamorphically similar to those described by Petrina in 1993. However, sporulation was no longer apparent, most likely due to prolonged storage and retransmissions. In at least one case, after five weeks of cultivation on oat agar, a mature perithecia was formed with the same sacs and ascospores as can be observed in nature. After incubation of the ascospores on agar formed from malt fermentation extract at room temperature, they germinated overnight. The mycelium was formed by highly branched, separated hyaline hyphae.

Sordarin is obtained by culturing a strain of Rosellinia subiculata or an unidentified fungus MF6232 (ATCC74387) capable of producing said compound on a conventional solid medium or in conventional liquid media. The organism is grown in a nutrient medium which contains known nutrient sources for similar fungi, i.e. assimilable carbon and nitrogen sources and optionally inorganic salts and other known growth agents. In the cultures of the present invention, it is possible to apply commonly used methods for culturing similar fungi.

0000

The nutrient medium should contain a suitable assimilable carbon source such as ribose, glucose, sucrose, cellobiose or fructose. The nitrogen source, which may be ammonium chloride, ammonium sulfate, urea, ammonium nitrate, sodium nitrate, etc., may be used either alone or in combination with organic nitrogen sources such as peptone, fish meal extract, yeast extract, "corn steep liquor," , soybean powder, cottonseed flour, etc.

If necessary, inorganic nutrient salts can also be added to the medium as sources of sodium, potassium, calcium, ammonium cations, phosphate anions, sulfate anions, chloride anions, bromide anions, carbonate anions, zinc, magnesium, manganese, cobalt, iron and the like.

Sordarin can be obtained at any temperature suitable for the satisfactory growth of the production organism, e.g. at a temperature of 20 to 30 ° C. Usually, optimal production of the desired compound is achieved in shake flasks after 7 to 21 days of incubation. Aeration in shake flasks is achieved by shaking, eg by shaking on a rotary shaker. If the fermentation is to be carried out in tank fermenters, it is desirable in the nutrient medium by inoculation of this nutrient medium with an oblique culture, lyophilized.

Even prepare a vegetative inoculum by culture or frozen culture.

After obtaining the active inoculum as described, this inoculum is transferred under aseptic conditions to the medium in the fermentation tank. Production of the desired compound in tank. fermenters is usually reached optimally after 7 to 21 days of incubation. The mixing of the mixture in the tank fermenter is performed by stirring and aeration can be performed by introducing air or oxygen into the stirred mixture. Compound production can be monitored by chromatographic or spectroscopic methods or by a conventional biological assay.

• · • · • ·

• «· • · ·

Sordarin is readily isolated from fermentation broth by extraction of the raw broth with an organic solvent such as methyl ethyl ketone. The obtained compounds can be purified by standard well-known methods such as gel filtration chromatography, thin layer chromatography, high performance liquid chromatography (HPLC), concentration, precipitation and / or crystallization, or combinations of these methods. Alternatively, the raw broth or its organic extract can be spray-dried or lyophilized and further purified by the above procedures.

The compound of the present invention (compound of formula I) can be produced by the following methods. These conditions are illustrative only and do not limit the scope of the invention in any way.

As shown in Scheme 1, a compound of formula (I) wherein Z is (a) or (b) may be prepared from a starting compound described in PCT application WO96 / 14326 or, in the case of compounds of formula (I) wherein Z is ( c), from the starting compound described in PCT application WO96 / 14327. The carboxyl group in the starting compound is first protected by a suitable protecting group (generally referred to as PG in the schemes), i. e.g.

benzyl or p-methoxybenzyl, followed by reaction with hydroxylamine hydrochloride in an alcoholic solvent containing pyridine to form an aldoxime. The resulting aldoxime is further converted with a suitable dehydrating agent (i.e., inner salt (methoxycarbonylsulfamoyl) triethylammonium hydroxide or cyanuric acid chloride). to the nitrile and deprotection affords the compound of formula (I).

• 0

Scheme 1

(AND)

P / OH is a lower alcohol solvent;

PG is a carboxyl protecting group;

Z is a group as defined above or a suitably protected derivative thereof

Scheme 2 shows the preparation of a compound of formula I (d). Sordarin is protected with a suitable protecting group and the aldehyde group is converted to the corresponding aldoxime by reaction with hydroxylamine hydrochloride in an alcoholic solvent containing pyridine, which is subsequently dehydrated with a dehydrating agent such as (methoxycarbonylsulfamoyl) triethylammonium hydroxide inner salt to give the nitrile compound (IV). Deprotection affords the compound of formula I (d).

9 9 9 • · • · 9 9 9 9

9 9 9 9 • 9 9 9 9

9 99999 9

9 9 9 9

99 99 999

Scheme 2 co ₂ pg

(IV)

Scheme 3 shows the microbial demethylation of the 4'-methoxy group in compound I (d) to give compound (V). Demethylation is achieved by contacting a compound of formula I (d) with a culture of a strain of Streptomyces avermitilis in a fermentation medium containing assimilable carbon and nitrogen sources; and isolating the compound of formula (V) from said fermentation medium. A group of suitable Streptomyces avermitilis strains includes strain MA4848 deposited with the American Type Culture Collection, Rockville, MD under the designation ATCC 31272. The compound of formula (V) may be used in the preparation of a compound of formula (I).

99 9 • 000 · · 9 9 9 9 9 • 99 9 9 0 0 0 0 9 99 9 • 999999 9 999 99 9 • · 9 ·· 9 9 9 9 9 ·· 9 9 9 · 0 0 0 0 0 00

Scheme 3 every ₂ h

microbial demethylation

(and)

As further shown in Scheme 4, sordaricin of formula (VI) can be used as a starting compound for the synthesis of compounds of formula (I). Protection of the carboxyl group with a suitable protecting group and subsequent protection of the primary hydroxyl group allows the synthesis of the nitrile-aglycone compound of formula (VII) by reaction with hydroxylamine hydrochloride, followed by dehydration of the resulting product and removal of the hydroxyl protecting group. Coupling of a suitable saccharide or modified saccharide substrate by well known methods provides a compound of formula (I).

• · · 1 • · • ··· * «• · • · • ·

(VI) (I)

The compounds of formula (I) are fungicidal agents which are useful in the manufacture of medicaments for humans and other animals. The compounds of formula (I) are also useful as crop protection agents.

The compounds of formula (I) are very active fungicides useful in the control of fungal infections in animals, including humans. They can be used, for example, in the treatment of fungal infections caused by organisms such as various species of the Candida strain (e.g. Candida albicans, Candida glabrata, (Torulopsis glabrata) _, Candida tropicalis and Candida pseudotropicalis), Cryptococcus neoformans, Pneumocystis carinii, Sp. (e.g. Aspergillus flavus and Aspergillus Coccidioides (Coccidioides immitis), Para (e.g. Paracoccidioides brasiliensis), HistoHistoplasma capsulatum) or Blastomyces (e.g.

Blastomyces dermatitidis). These compounds can also be used to treat other fungal infections caused by species

Aspergillus fumigatus), plasma coccidioides (eg ·· ·· ·· ♦ · fe · · • fefefe · • ··· Trichophyton, Microsporum or Epidermophyton strains (eg Trichophyton mentographytes, Trichophyton rubrum, Microsporum canis or Epidermophyton floccosum) or for treatment mucosal infections caused by Candida albicans.

The compounds of formula (I) may further be used to treat other infections caused by various species of filamentous fungi such as Geotrichum (e.g. Geotrichum clavatum), Trichosporon (e.g. Trichosporon beigelii), Blastoschizomyces (e.g. Blastoschizomyces capitatus), Sporothrix (e.g. Sporothrix) schenckii), Scedosporium (e.g. Scedosporium apiosperum), Cladosporium (e.g. Cladosporium carrionii) and Pityrosporum oval.

The compounds of formula (I) may also be used to treat infections caused by protozoa such as Toxoplasma, Cryptosporidium, Leishmania, Tripanosoma, Giardia and Trichomonas.

Determination of the in vitro fungicidal activity of the compounds of the present invention was performed on a liquid or solid medium by a fungicidal two-fold serial dilution method to determine the minimum inhibitory concentration (MIC) of the fungicidal agent at which growth inhibited after 24 to 48 hours of incubation at 35 ° C. In practice, the assay was performed by inoculating a series of agar plates or microdilution broth panels containing doubly diluted test fungicide with a standard culture of a clinically relevant pathogen such as Candida albicans. Then, agar plates or microdilution broth panels were tested for or not mold growth and the appropriate minimum inhibitory concentrations (MICs) were recorded. A live strain of Almar Blue was used to visualize the endpoints.

• · • · · · · · • · · · · · · · · · ····· ·

Determination of the in vivo fungicidal activity of the compounds of formula (I) can be performed by a series of different doses administered (e.g. subcutaneously, orally, intraperitoneally or intravenously) to mice that have been inoculated intravenously with Candida spp. The kidneys can be removed from the test animals and the content of a viable strain of Candida spp. and a reduction in infection can be determined relative to an untreated control group of animals.

In terms of fungicidal activity, the compounds of formula (I) are useful for the treatment and / or prevention of various fungal diseases in humans and other animals. Such infections include superficial, skin, subcutaneous and systemic fungal infections such as respiratory tract infections, gastrointestinal infections, cardiovascular infections, urinary tract infections, central nervous system infections, candidiasis and chronic mucocandidosis (e.g. aphthous candidiasis and vaginal candidiasis) and skin infections caused by fungal, cutaneous and mucosal candidiasis, dermatophytosis including ring lichen and infections dermatophytosis of the tina, dermatophytosis of the feet, inflammation of the nail bed, pityriasis versicolor, erythrasma, sores, fungal rash, candida vulvoduis and candida balavitis. Furthermore, the compounds of general formula (I) can be used as prophylactic agents for the prevention of systemic and topical fungal infections. The use of these compounds as prophylactic agents may be useful, for example, in a diet for selective decontamination of the gut in preventing infection in immunocompromised patients (e.g., AIDS patients, cancer patients, or transplant patients). In the case of some disease syndromes or iatrogenic conditions, it may also be desirable to prevent excessive fungal growth during antibiotic treatment.

• 999

9 • · 999 · 9

The compounds of formula (I) also have a wide range of uses as crop protection agents and are effective against a wide range of phytopathogenic fungi, in particular fungi from the group comprising deuteromycetes (eg Botritis spp., Septoria spp., Pyricularia spp., Stagnospora spp. ., Helminthosporium spp., Fusarium spp., Cercospora spp., Rynchosporium spp., Pseudocercosporella spp. And Alternaria spp.); basidiomycetes (e.g. Puccinia spp., Rhizoctonia spp. and Hemileia); ascomycetes (e.g., Venturia spp., Podospharera spp., Erysiphe spp., Monilinia spp. and Uncinula spp.); and oomycetes (e.g., Phytophthora spp., Pemospora spp., Bremia spp., Pythium spp. and Plasmopara spp.). The foregoing list gives examples of phytopathogenic fungi against which the compounds of the present invention exhibit fungicidal effects and is not limiting in any sense. The compounds of the present invention have very advantageous medicinal and preventive fungicidal properties suitable for plant protection and can be used for inhibiting or killing microorganisms occurring on plants or parts of plants (fruits, flowers, leaves, stems, tubers or roots) or on various crops. useful plants, while at the same time those parts of the plants which grow later are also protected against such microorganisms. The compounds according to the invention can also be used as additives in the treatment of plant growth material, in particular seeds (fruits, tubers, grains) and plant cuttings (for example rice) to provide protection against fungal infections and against phytopathogenic fungi occurring in the soil. The compounds of the formula (I) according to the invention are characterized by the fact that they are particularly well tolerated by plants and, in addition, are environmentally friendly.

ft * ft * * · ftftftft ftft ftft ftftftft ftftft ftftftft ftftftft ftft ··· ft ftft · • ··· ftft · ftftft ftft · • ft · ftft ft ftftftft ftftftft ftft ftft ftft ftft

The determination of the agricultural utility of the compounds of formula (I) can be carried out by the following tests:

1. Efficacy against Erypsiphe gramínis on wheat

a) After culturing for one week, the wheat plants were treated by spraying containing 200 ppm of active ingredient, 20% of acetone and 0.25% of Triton X155 until it was consumed. After 2 hours, the treated plants were infected with ascospores shaken from the inoculum. To determine the degree of protection provided by the test compound, fungal infestation was evaluated after eight days of incubation at 22 ° C and 50% relative humidity.

b) After culturing for one week, the wheat plants were infected with ascospores shaken from the inoculum. After 24 hours, the wheat plants were treated with a spray containing 200 ppm of active ingredient, 20% acetone and 0.25% Triton X155. To determine the therapeutic effect provided by the test compound, fungal infestation was evaluated after an eight-day incubation at 22 ° C and 50% relative humidity.

c) After culturing for one week, the wheat plants were infected with ascospores shaken from the inoculum. After 24 hours, the soil in which the plants were grown was watered with a dressing containing 200 ppm of active ingredient, 20% acetone and 0.25% Triton X155. To determine the therapeutic effect provided by the test compound, fungal infestation was evaluated after an eight-day incubation at 22 ° C and 50% relative humidity.

2. Efficacy against Puccinia recondita on wheat

a) After one week of cultivation, the wheat plants were treated with a spray containing 200 ppm of active ingredient, • 444 · · 4 4 4 4 4 • 444 44 444 4 444

444444 4 444 44 4

4 44 4 4444

44 44 444 44 44% acetone and 0.25% Triton X155, until consumed. After 2 hours, the treated plants were infected with spores. To determine the degree of protection provided by the test compound, fungal infestation was evaluated after one day of incubation at 20 ° C and 95-100% relative humidity followed by a seven-day incubation at 25 ° C and 50% relative humidity.

b) After culturing for one week, the wheat plants were infected with a spore-containing suspension. After 24 hours, the wheat plants were treated by spraying containing 200 ppm of active ingredient, 20% of acetone and 0.25% of Triton X155 until consumed. To determine the therapeutic effect provided by the test compound, fungal infestation was evaluated after one day of incubation at 20 ° C and 95-100% relative humidity followed by a seven-day incubation at 25 ° C and 50% relative humidity.

c) After culturing for one week, the wheat plants were infected with a spore-containing suspension. After 24 hours, the soil in which the plants were grown was watered with a dressing containing 200 ppm of active ingredient, 20% acetone and 0.25% Triton X155. To determine the therapeutic effect provided by the test compound, fungal infestation was evaluated after one day of incubation at 20 ° C and 95-100% relative humidity followed by a seven-day incubation at 25 ° C and 50% relative humidity.

Based on the extent of the effects, the compounds according to the invention can be used for the protection or treatment of plants against infestation or already infested with phytogenic fungi which infest various agricultural crops. The following list includes those plant species which are suitable for the applications of the compounds of the invention described in the present invention: cereals (wheat, rye, oats, barley, rice, sorghum and related ·· φ * Φ · φφφφ · φ · * • φφφ φφφ φ φ φ «• · · · · · ΦΦΦ · 9 9 9

9 999 9 9 Φ ΦΦΦ 9 9 Φ • · Φ ΦΦ Φ ΦΦΦΦ ·· ·· ΦΦ ΦΦΦ ΦΦ φφ crops); beet (sugar beet or fodder beet); mallow, stone and soft fruits (eg apples, pears, plums, peaches, almonds, cherries, strawberries, raspberries and blackberries); legumes (eg beans, peas, lentils and soybeans); oilseeds (rape, mustard, poppy, olives, sunflowers, coconut, castor oil producing plants, cocoa beans and groundnuts); pumpkin plants (eg cucumber, squash and watermelon); textile plants (eg cotton, flax, hemp and jute); citrus fruits (eg oranges, lemons, tangerines and grapefruits); vegetables (eg lettuce, cabbage, spinach, carrots, asparagus, peppers, onions, tomatoes and potatoes); plants of the laurel family (avocado, cinnamon and camphor); or plants such as corn, tobacco, nuts, coffee, sugar cane, tea, vines, hops, bananas and rubber trees as well as ornamental plants (flowers, shrubs, deciduous trees and evergreen plants such as conifers). However, not all of these plant species constitute a limiting list of plants with respect to the spectrum of activity of the compounds of this invention.

The compounds of general formula (I) are particularly suitable for the treatment of the following plant diseases:

Erysiphe graminis in cereals, Erysiphe cichoracearum and Sphaerotheca fuliginea in pumpkins, Podasphaera leucotricha in apples, Uncinula necator in vines, Puccinia. sp. in cereals, Rhizoctonia solani in cotton, Ustilago sp. for cereals and sugar cane, Venturia inaequalis (scab) for apples, Helminthosporíum sp. in cereals, Septoria nodorum in wheat, Botrytis cinerea (gray form) in strawberries and grapes, Cercospora arachidicola in peanuts, Pseudocercosporella herpotrichoides in wheat and barley, Pyricularia oryzae in rice, Phytophtora infestans in potatoes and tomatoes, Fus. and Verticillium sp. in various plants,

0000

Plasmopara viticola

04 • · · · · • · 4 0 · 000 0 ·

0 00

04 * 4 • 044 • 0 004

0 00

0 · 1

0 4 (• 0 0 0 1

0 4 4

04 grapes, Alternaria sp.

fruit vegetables. The compounds of formula (I) may also be used to protect various materials (eg to protect wood raw material from Paecilomyces variotii).

Pharmaceutical products

Although it is in principle possible for the compounds according to the invention to be administered therapeutically on their own, i.e. as raw material, it is preferred to administer the active ingredient in the form of a pharmaceutical composition. Therefore, the invention further relates to a pharmaceutical composition, a compound of formula (I), or which comprises a pharmaceutically acceptable salt of a compound of formula (I), together with one optionally acceptable other or more pharmaceutically acceptable carrier of a compound of formula (I) and therapeutic and / or prophylactic ingredients.

The carrier (s) must be acceptable in the sense of being compatible with the other ingredients of the pharmaceutical composition and not deleterious to the recipient thereof.

The pharmaceutical compositions of the present invention include pharmaceutical compositions for oral, buccal, parenteral, implantation, rectal, topical, ophthalmic or genitourinary administration, or in a form suitable for administration in the form of implants or in a form suitable for inhalation or insufflation administration.

Tablets and capsules for oral administration may contain conventional excipients such as binding agents, for example syrup, acacia, gelatin, sorbitol, tragacanth, starch glue or polyvinylpyrrolidone; pincers, for example lactose, sugar, microcrystalline cellulose, corn starch, calcium phosphate or sorbitol; lubricants, eg magnesium stearate, gourds29 ·· 4 · 4444 44 44 • 444 4 4 4 4 4 4 4 • · 4 · · · 444 · 4 4 4

4 444 4 4 4 444 4 4 4

4 44 4 4444

44 4 · 444 44 44 Stearic line, talc, polyethylene glycol or silica; disintegrants, for example potato starch or sodium starch glycolate or sodium cross-caramel; or wetting agents such as sodium lauryl sulfate. Tablets, which include chewable, soluble and effervescent tablets, may be coated according to well known methods. Oral liquid preparations may be in the form of, for example, aqueous or oily suspensions, solutions, emulsions, syrups or elixirs, or may be presented as a dry product for constitution with water or other suitable vehicle just prior to use. Such liquid pharmaceutical compositions may contain conventional additives such as suspending agents, for example sorbitol syrup, methylcellulose, glucose / sugar syrup, gelatin, hydroxyethylcellulose, carboxymethylcellulose, aluminum stearate gel or hardened fats; emulsifying agents, for example lecithin, sorbate monooleate or acacia; non-aqueous vehicles (which may include edible oils), for example almond oil, fractionated coconut oil, oily esters, propylene glycol or ethyl alcohol; and preservatives, for example methyl or propyl p-hydroxybenzoates or sorbic acid.

The pharmaceutical composition of the present invention for buccal administration may be in the form of tablets or lozenges formulated by conventional means.

The pharmaceutical composition of the present invention for parenteral administration may be formulated for administration by injection or continuous infusion. Pharmaceutical compositions for injection may be presented in unit dosage form in ampoules or in multi-dose bottles, with an added preservative. These pharmaceutical compositions may take such forms as suspensions, solutions, or emulsions in oily or oily materials

9 9 9 9

9 9 ·· · *

9 9 9 • 9 99

9

Aqueous vehicles and may contain formulatory agents such as suspending agents, stabilizing agents and / or dispersing agents. Furthermore, the active ingredient may be in powder form for constitution with a suitable vehicle, e.g., pyrogen-free water, before use.

For inhalation administration, they are easily administered in the form of an aerosol spray from pressurized packs using a suitable propellant gas, e.g. dichlorodifluoromethane, trichlorofluoromethane, dichlorotetrafluoroethane, carbon dioxide or other suitable gases, or from a nebulizer. In the case of the use of a pressurized aerosol, it is possible to determine the individual dose by allowing the valve to deliver a metered amount of aerosol.

Another possibility for inhaled administration of a pharmaceutical composition of the invention is that the pharmaceutical composition is in the form of a dry powder composition, for example a powder mixture of a compound of the invention and a suitable powder base such as lactose or starch, or modified physical forms of the drug substance itself. The powder composition may be presented in unit dosage form, e.g., in capsules or cartridges of, e.g., gelatin, or in blister packs from which the powder may be administered by means of an inhaler or insufflator.

The pharmaceutical composition of the present invention may be in the form of a suppository comprising, for example, a conventional suppository base, or the composition may be in the form of a pessary comprising, for example, a conventional pessary base.

The pharmaceutical compositions of this invention may also be formulated for topical administration in the form of ointments, creams, gels, lotions, shampoos, powders (including spray powders), pessaries, tampons, sprays, therapeutic baths, aerosols, aerosols, 0 00 0000 * 00 0004

00 0 0 0000 0 00 1 0 0000 * 0 0 * * * 0 * «00 0 00 0 000«

The vehicle may comprise drops (eg eye drops, ear drops or nasal drops) or overflow solutions. Ointments and creams may, for example, be formulated with an aqueous or oily base with the addition of suitable thickening or gelling agents. Ointments for administration to the eye may be manufactured under sterile conditions using sterile ingredients. Overflow solutions can be formulated, for example, for veterinary use in organic solvents containing oils, optionally formulation agents such as stabilizers and solubilizers. Pessaries and tampons for insertion into the vagina may be formulated in conventional manner and, where appropriate, may contain effervescent. Such pharmaceutical compositions may also contain other active ingredients such as suitable corticosteroids, antibiotics or antiparasitics.

Liquid preparations for intranasal use may take the form of solutions or suspensions, and may include conventional vehicles such as tonicity agents, for example sodium chloride, dextrose or mannitol; preservatives, for example benzalkonium chloride, thiomersal, phenylethyl alcohol; and other formulation agents such as suspending agents, buffering agents, stabilizing agents, dispersing agents, and flavoring agents.

Transdermal administration may be effected by providing a suitable system which promotes the absorption of the active compound through the skin and generally consists of a base composition enclosed in an adhesive patch comprising carrier films, membranes and release binders. Such systems may include absorption enhancers such as alcohols or may function to promote iontophoresis.

The pharmaceutical composition of the present invention may also be formulated as a depot preparation. Such a long32 • · · »· · · • · · · • · 9 • ·« «· ·

the agents may be administered by implantation (for example under the skin or into a muscle) or by intramuscular injection. Thus, for example, a compound of the present invention may be formulated with suitable polymeric or hydrophobic materials (for example as an emulsion in a pharmaceutically acceptable oil) or ion exchange resins, or as almost insoluble derivatives, e.g., as an almost insoluble salt.

When the pharmaceutical composition of the present invention contains individual doses, each dose preferably contains from 0.001 milligram to 1000 milligrams, preferably from 0.01 milligrams to 400 milligrams, of active ingredient in cases where the compound of the present invention is to be administered orally. The daily dose used for the treatment of adults is preferably from 0.001 milligram to 5000 milligrams of active ingredient, more preferably from 0.01 milligrams to 2000 milligrams of active ingredient. This dose can be administered, for example, in 1 to 4 doses, depending on the route of administration and the condition of the patient and the disease to be treated.

The compound of the invention may be administered, for example, by intravenous infusion at a dose of up to 50 milligrams of active ingredient / kilogram / day. The duration of treatment is determined by the rate of response rather than by a predetermined number of days.

The compounds of the invention may also be used in combination with other therapeutic agents, and therefore combinations of the compound of the invention together with another therapeutically active agent are a further aspect of the invention.

Thus, for example, the compounds of the present invention may be used in combination with one or more fungicides, such as a polyene derivative (e.g., amphotericin B, nystatin, a lipid formulation of amphotericin B); azole derivative (eg fluconazole, ftft • ftft ftft ftft ftft ftft ftft ftft ftft ftft ftft ftft ftft ftft ftft ftft ftft ftft ftft ftft ftft ftft ftft ftft ftft ftft ftft ftft ftft ftft ftft ftft ftft ftft ftft ftft ftft ftft ftft ftft ftft ftft ftft ftft ftft ftft ftft ftft ftft ftft ftft ftft ftft ftft ftft ftft ftft ftft ftft ftft ftft ftft ftft ftft ftft Itraconazole, ketoconazole, miconazole, clotrimazole, ZD-08070, UK-109496, SCH56592), 5-fluorocytosine, a pneumocandin or echinocandin derivative such as cilofungin, LY-303366, L- 733560, L-743872 or other cell wall compounds such as nikcomycin Z and / or one or more immunodeficiency compounds such as interferon e.g. (IFN-), interleukin e.g. (IL-1, IL-2, IL-3 and IL-8) and cell stimulating agents, [(G) -CSF, (M) -CSF and (GM) -CSF] and defensins. Particularly suitable compounds for use with the compounds of the present invention are itraconazole, flucytosine, fluconazole or amphotericin B.

When the compounds of the present invention are administered in combination with another fungicidal compound, the compound of the present invention and the other fungicide may be administered at the recommended maximum clinical doses or at lower doses.

The above combinations may be intended for use in the form of pharmaceutical compositions, and therefore pharmaceutical compositions comprising the above-described combinations together with a pharmaceutically acceptable carrier for this combination are a further aspect of the invention. The separate components of such a pharmaceutical composition may be administered either sequentially or simultaneously in separate or combined pharmaceutical compositions.

When a compound of the invention is used in combination with a second drug for the same indication, the dosages of the individual compounds may differ from the case where each of the compounds is used alone. Suitable doses easily. determined by an expert in the field.

• · · «• · • · ·· • ·

Agrochemical means

The compounds of general formula (I) can be used either in unchanged form or preferably together with excipients which are commonly used in the field of agrochemicals and are known for their use mainly as: emulsifiable concentrates, paintable pastes, directly sprayable or dilutable solutions, dilute solutions, suspensions (including aqueous, oily or other suspensions with a high percentage of water), dispersions, oily dispersions, spreading agents, wetting powders, soluble powders, dusts, granules and encapsulated compositions. The agrochemical compositions according to the invention are prepared by known methods, for example by homogeneously mixing and / or grinding the active ingredients with additional ingredients, for example solvents, solid carriers and, where appropriate, surfactants. Powders, dusts and spreading agents can be prepared by mixing or grinding the active ingredients with a solid carrier. Granules, for example coated granules, impregnated granules or homogeneous granules, can be prepared by binding the active ingredients to solid carriers.

A group of suitable solvents includes: aromatic hydrocarbons, preferably fractions containing 8 to 12 carbon atoms, such as mixtures of xylenes or substituted naphthalenes, chlorinated aromatics such as chlorobenzenes, phthalates such as dibutyl or dioctyl phthalate, aliphatic hydrocarbons such as cyclohexane or paraffins, alcohols and glycols and their ethers and esters, such as ethanol, ethylene glycol, ethylene glycol monomethyl ether or ethylene glycol monoethyl ether, ketones, such as cyclohexanone, amines, such as ethanolamine, strongly polar solvents, such as N-methyl-2-pyrrolidone, dimethyl sulfoxide or N, N- dimethylformamide and vegetable oils or epoxidized

9 • ·· · • ·

vegetable oils such as epoxidized coconut oil or soybean oil and water.

Examples of surfactants are: salts of aromatic sulfonic acids containing alkali metals, alkaline earth metals and ammonium cations, e.g. ligninsulfonic acid, phenolsulfonic acid, naphthalenesulfonic acid and dibutylnaphthalenesulfonic acid and the same fatty acid salts, alkyl and alkylarylsulfonates and alkyl lauryl ether and sulphates of fatty alcohols and salts of sulphonated hexadecanols, heptadecanols and octadecanols, salts of glycol ethers of fatty alcohols, products of condensation of sulphonated naphthalene derivatives with formaldehyde, condensation products of naphthalenes or naphthalenesulphonic acid derivatives with phenol and formaldehyde ethoxyolytolephenolphenolphenolphenolphenyl alkylaryl polyether alcohols, isotridecyl alcohol, condensates of fatty alcohols with ethylene oxide, ethoxylated castor oil, polyoxyethylene alkyl ethers, ethoxylated polyoxypropyl ethylene, acetal lauryl alcohol polyglycol ether, sorbitol esters, waste liquor of lignin sulfite and methylcellulose.

Examples of solid carriers are mineral soils such as silicic acids, silica gels, silicates, talc, kaolin, attapulgite, limestone, lime, chalk, trunk, loess, clay, dolomite, kieselguhr, aluminum-calcium sulphate, magnesium sulphate, magnesium oxide, ground plastics, fertilizers such as ammonium sulphate, ammonium phosphate, ammonium nitrate and urea and its derivatives and vegetable products such as wholemeal flour, ground bark or ground wood and ground walnut shells, cellulose powders, etc.

• · · · ·

The compounds of formula (I) can be mixed and applied

► · · <

»· · 1» · · 4 ·· ·· together with other active insecticides, bactericides, ingredients such as herbicides, nematicides, anti-mollusc agents, growth regulators, micronutrients and fertilizers. The other ingredients may also include one or more fungicides belonging to, but not limited to, the following classes of fungicides: carboxamides, benzimidoazoles, triazoles, hydroxypyridines, dicarboxamides, phenylamides, thiadiazoles, carbamates, cyanooximes, cinnamic acid derivatives, morpholines, imidazoles, and B-methoxyacrylates pyridines / pyrimidines. Furthermore, these other active ingredients may be used as mixtures of several agrochemical compositions of the present invention and, if desired, together with other excipients commonly used in the art to facilitate the application of the composition. Suitable carriers or excipients may be solid or liquid and correspond to substances which are generally used in the processes for making these compositions or regenerated mineral compounds, solvents, dispersants and wetting agents).

The following list of fungicides with which the compounds of formula (I) may be combined is given only to illustrate possible combinations without any limitation with respect to other possible combinations. Examples of fungicides which can be combined with the compounds of formula (I) are: sulfur, dithiocarbamates and their derivatives such as ferric dimethyldithiocarbamate, zinc dimethyldithiocarbamate, zinc ethylenebisdithiocarbamate, manganese-containing ethylenebisdithiocarbamate thiocarbamate thiocarbonate, manganese and , Zinc N'-ethylenebisdithiocarbamate, zinc ammonium complex N, N'-propylenebisdithiocarbamate, technological N, N'-propylenebisbate (eg natural • 4 · · · 4 · 4 · · · *

44 4 4 4444 4 44 4

4 444 44 4 44 4-44 4

44 44444 44 44 Zinc dithiocarbamate, N, N'-polypropylene bis (thiocarbamyl) disulfide; nitro derivatives such as dinitro (1-methylheptyl) phenyl crotonate, 2-sec-butyl-4,6-dinitrophenyl-3,3-dimethylacrylate, 2-sec-butyl-4,6-dinitrophenyl isopropyl carbonate and diisopropyl 5-nitroisophthalate; heterocyclic compounds such as 2-heptadecylimidazol-2-ylacetate, 2,4-dichloro-6- (o-chloroanilino) -s-triazine, O, O-diethylphthalimidophosphonothioate, 5-amino-1- [bis (dimethylamino) phosphinyl] - 3-phenyl-1,2,4-triazole, 2,3-dicyano-1,4-dithioanthraquinone, 2-thio-1,3-dithio [4,5-b] quinoxaline, methyl-1- (butylcarbamoyl) - 2-benzimidazole carbamate, 2-methoxycarbonylaminobenzimidazole, 2- (fur-2-yl) benzimidazole, 2- (thiazol-4-yl) benzimidazole, N- (1,1,2,2-tetrachloroethylthio) tetrahydrophthalimide, N-trichloromethylthiotetrahydrophthalimide, N -trichloromethylthiophthalimide, N-dichlorofluoromethylthio-N ', N'-dimethyl-N-phenylsulfondiamide, 5-ethoxy-3-trichloromethyl-1,2,3-thiadiazole, 2-thiocyanomethylthiobenzthiazole, 1,4-dichloro-2,5-methoxybenzene, 4 - (2-chlorophenylhydrazono) -3-methyl-5-isoxazolone, 2-thiopyridine-1-oxide, 8-hydroxyquinoline and its copper-containing salt, 2,3-dihydro-5-carboxanilido-6-methyl-1,4-oxathiine, 2,3-dihydro-5-carboxanilido-6-methyl-1,4-oxathiine-4,4-dioxide, 2-methyl-5,6-dihydro-4H-pyran-3-carboxanilide, 2-methylfuran -3-carboxanilide, 2,5-dimethylfuran-3-carboxanilide, 2,4,5-trimethylfuran-3-carboxanilide, 2,5-dimethyl-N-cyclohexylfuran-3-carboxanilide, N-cyclohexyl-N-methoxy-2 , 5-diethyl-furan-3-carboxanilide, 2-methylbenzanilide, 2-iodobenzanilide, N-formyl-N-morpholine-2,2,2-trichloroethyl acetal, piperazine-1,4-diylbis (1- (2,2,2- trichloroethyl) formamide),

1- (3,4-dichloroanilino) -1-formylamino-2,2,2-trichloroethane,

2,6-dimethyl-N-tridecylmorpholine and its salts, 2,6-dimethyl-N-cyclododecylmorpholine and its salts, N- [3- (p-tert-butylphenyl) -2-methylpropyl] -cis-2,6-dimethyl- morpholine, N- [3- (p-tert-butyl38 phenyl) -2-methylpropyl] piperidine, 1- [2- (2,4-dichlorophenyl) -4-ethyl-1,3-dioxolan-2-yl-ethyl ] -1H-1,2,4-triazole, 1- [2- (2,4-dichlorophenyl) -4-n-propyl-1,3-dioxolan-2-yl-ethyl] -1H-1,2,4-triazole, N- (n-propyl) -N- (2,4, β-trichlorophenoxyethyl) -N-imidazoleurea, 1- (4-chlorophenoxy) -3,3-dimethyl-1- (1H-1,2,4-triazole -1-yl) butan-2-one, 1- (4-chlorophenoxy) -3,3-dimethyl-1- (1H-1,2,4-triazol-1-yl) butan-2-ol, α- (2 chlorophenyl) -α- (4-chlorophenyl) -5-pyrimidine-methanol, 5-butyl- (2-dimethylamino-4-hydroxy-6-methylpyrimidine, bis (p-chlorophenyl) -3-pyridinemethanol,

1,2-bis (3-ethoxycarbonyl-2-thioureido) benzene, 1,2-bis (3-methoxycarbonyl-2-thioureido) benzene and various fungicides such as dodecylguanidine acetate, 3- [3- (3,5-dimethyl- 2-oxycyclohexyl) -2-hydroxyethyl] glutaramide, hexachlorobenzene, DL-methyl-N- (2,6-dimethylphenyl) -N- (fur-2-yl) alanate, methyl DL-N- (2,6-dimethylphenyl) -N - (2-methoxyacetyl) alanate, N- (2β-dimethylphenyl) -N-chloroacetyl-DL-2-aminobutyrolactone, methyl DL-N- (2,6-dimethylphenyl) -N- (phenylacetyl) alanate, 5-methyl- 5-vinyl-3- (3,5-dichlorophenyl) -2,4-dioxo-1,3-oxazolidine, 3- [3,5-dichlorophenyl] -5-methyl-5-methoxymethyl-1,3-oxazolidine-2, 4-dione, 3- (3,5-dichlorophenyl) -1-isopropylcarbamoylhydantoin, N- (3,5-dichlorophenyl) -1,2-dimethylcyclopropane-1,2-dicarboximide, 2-cyano- [N (ethylaminocarbonyl) - 2-methoximino] acetamide, 1- [2- (2,4-dichlorophenyl) pentyl] -1H-1,2,4-triazole, 2,4-difluoro-α- (1H-1,2,4-triazole-1- ylmethyl) benzhydryl alcohol, N- (3-chloro-2,6-dinitro-4-trifluoromethylphenyl) -5-trifluoromethyl-3-chloro-2-aminopyridine and 1 - ((bis (4-fluorophenyl) methylsilyl) methyl-1H-1, 2,4-tr iazole.

Depending on the nature of the agrochemical composition of the present invention and the objects to which the composition is to be applied, and the prevailing conditions, the method of application such as spray spraying, fine spraying, spraying is selected.

0 · · »♦ · 0

I · · · ► · 0 0 0 «· * ·» »» 4 »· · <

»· 0 <

dispersing, coating, wrapping and watering. One method of applying an active ingredient or agrochemical composition comprising at least one of said compounds is to apply to plants (i.e., foliar application); However, the active ingredient may penetrate the plant roots through the soil (i.e. by soil application). This application can be carried out either by applying the liquid to the soil (by feeding) or by applying granules containing the active ingredient (s).

The active ingredient according to the invention can also be applied to plant growth material, such as seeds (fruits, tubers or grains) or plant cuttings, either in liquid form (coating) or in solid form (coating). For example, the seeds may be coated before sowing. The compounds of the present invention can also be applied to grains, either by impregnating the grains in a liquid form or by coating the grains with a solid form of the agrochemical composition of the present invention. The agrochemical can also be applied directly to the place where the plants are grown, for example to the seed furrows during sowing.

The usual preferred amount of composition is from 10 grams to 50 kilograms of active ingredient per hectare, preferably 100 grams to 2 kilograms of active ingredient per hectare, more preferably 100 to 600 grams per hectare. The active ingredients of said compounds are generally used in the form of agrochemicals and can be applied to plants or plant parts either simultaneously or in sequence with the other active ingredients. These other active ingredients may be fertilizers, other micronutrients or other compounds affecting plant growth. However, they can also be selective herbicides, insecticides, bactericides, nematicides and anti-mollusc agents as well as other fungicides.

• 9 99 99 9999

9999 999 9999

99 9 9 9999 9 99 ·

999999 9 999 99 9

9 99 9 9999

99 99 999 99 99

Preparation of starting material

Fermentation production of sordarin

For the cultivation of Rosellinia subiculata (ATCC 74386) a

ATCC 74387, the following media were used in the production of sordarin:

Vaccination medium 1

Component

Yeast extract Malt extract Glucose

Junlon gram / liter

4.0

8.0

4.0

1.5

The medium was prepared in distilled water, the pH was adjusted to 7.0 before sterilization, and then the medium was dispensed in 50 milliliters into uncovered 250 milliliter Erlenmeyer flasks. The flasks were closed with cotton plugs and subsequent sterilization was performed at 121 ° C for 20 minutes.

Vaccination medium 2

Gram / liter component

Maize extract (dried) 2.5

Tomato paste 40.0

Oatmeal 10.0

Glucose 10.0

Trace element solution 10.0 ml / l

·· • fefefe fe fefe fe fe • · • ·

Trace element solution

Component

FeSO ₄ .7 H ₂ 0 MnSO ₄ .4 H ₂ 0 CuCl ₂ .2 H ₂ 0 CaCl ₂ .H ₂ 0

H3BO3 (NH ₄ ) _with Mo0 ₂₄ .4 H ₂ 0 ZnSO ₄ .7 H ₂ 0 gram / liter

1.0

1.0

0.025

0.1

0.056

0.019

0.2

A solution of trace elements was prepared in 0.6 N hydrochloric acid

The medium was prepared in distilled water, the pH was adjusted to 6.8 before sterilization, and then the medium was dispensed in 50 milliliters into uncovered 250 milliliter Erlenmeyer flasks. The flasks were closed with cotton plugs and subsequent sterilization was performed at 121 ° C for 20 minutes.

Solid production medium 1

1. Fixed part

675 milliliters of vermiculite were dispensed into a 2 liter cylindrical bottle. The bottle was closed with a latex stopper and placed in an autoclave for 60 minutes and then dried for 30 minutes.

2. Liquid part

220 milliliters of the following mixture were weighed into a 500 milliliter bottle:

0

0 0 0 • 00 0 0

0 0 0 • 0 • 000

00 «0 1

0 «

Component gram / liter Glucose 150.0 Glycerol 20.0 Yeast extract 4.0 NaNO ₃ 1.0 Sodium glutamate 3.0 « At ₂ HPO ₄ 0.5 * MgSO ₄ .7 H ₂ O 1.0 K-elements 1.0 ml / l CaCO ₃ 8.0 K-elements Component gram / liter FeCl ₃ .6 H ₂ O 5.8 MnSO ₄ .H ₂ O 0.1 CoC1 ₂ .6 H ₂ 0 0.02 CuSO ₄ .5 H ₂ O 0.015 Na ₂ Mo0 ₄ .2 H ₂ O 0.012 ZnCl ₂ 0.02 SnCl ₂ .2 H ₂ 0 0.005 AND h ₃ bo ₃ 0.01 KC1 0.02 AND HCl (concentrated) 2.0 ml / l

The medium was prepared in distilled water, the pH was adjusted to 7.0 before sterilization, the medium was divided into 500 ml bottles and then heat-treated in an autoclave for 15 minutes at 121 ° C. Glucose was heat treated in an autoclave separately from the other components.

·

0 * 0 4

44 4 4 ♦ · 4 ·

0 0 «

0 4 «• 4 4 4

4 «44

Liquid production medium 1

Component gram / liter Glycerol 75.0 Glucose 75, 0 Tomato paste 5.0 NZ amine type A 4.0 Ardamin PH 5.0 K ₂ HPG ₄ 0.5 MgSO ₄ .7 H ₂ O 0.25 KCI 0.25 ZnSO ₄ .7 H ₂ O 0.5 CaCO ₃ 10.0

The medium was prepared in distilled water, the pH was adjusted to 7.0 before sterilization, and the medium was dispensed in 50 ml portions into uncovered 250 ml Erlenmeyer flasks. The flasks were closed with cotton plugs and subsequent sterilization was performed at 121 ° C for 20 minutes.

Solid production medium 2

1. Fixed part

675 milliliters of vermiculite were dispensed into a 2 liter cylindrical bottle. The bottle was closed with a latex stopper and placed in an autoclave for 60 minutes and then dried for 30 minutes.

2. Liquid part

220 milliliters of the following mixture were weighed into a 500 milliliter bottle:

Μ 44 • 4 4 4 ·

4 4 4 4 • 4 · 44 4 4

4 4 · * 4 4 4 4 4 ♦ 44

44

4 ♦ 4

4 • 4

4

Component gram / liter Sucrose 60.0 Glucose 80.0 Glycerol 60.0 Citric acid 15, 0 NZ amine type A 5.0 NaNO ₃ 1.0 kh ₂ after ₄ 0.5 MgSCú. 7 H ₂ O 0.5 CaCO ₃ 0.5 K-elements 1 ml / l

K-elements

Component gram / liter FeCl ₃ .6 H ₂ 0 5.8 MnSO ₄ .H ₂ O 0.1 CoCl ₂ .6 H ₂ 0 0.02 CuSO ₄ .5 H ₂ O 0.015 Na ₂ Mo0 ₄ .2 H ₂ 0 0.012 ZnCl ₂ 0.02 SnCl ₂ .2 H ₂ O 0.005 h ₃ bo ₃ 0.01 KC1 0.02 HCl (concentrated) 2.0 ml / l

The medium was prepared in distilled water, the pH was adjusted to 1.0 before sterilization, the medium was dispensed in 220 milliliters into 500 milliliter bottles and heat-treated in an autoclave for 15 minutes at 121 ° C.

• · ·

»· * · ♦ ·

·· * · • · · ·

Liquid production medium 2

The composition was identical to the liquid part of the solid production medium 1. The medium was prepared in distilled water, the pH was adjusted to 7.0 before sterilization. The medium was dispensed in 50 milliliters portions into uncovered 250 milliliter Erlenmeyer flasks. The flasks were capped and heat treated in an autoclave at 121 ° C for 15 minutes. Glucose was heat treated in an autoclave separately from the other components.

Sordarin production by fermentation of Rosellina subiculata (MF6239, ATCC 74386)

1. Culture:

An oblique section of the culture-containing agar was aseptically transferred to inoculum medium 1 (50 milliliters in an uncovered 250 milliliter flask). To obtain biomass, the mixture was incubated on a swinging cone shaker (cone diameter was approximately 5.1 centimeters (2 inches)) for 5 days at 220 rpm, 25 ° C and 85% relative humidity. The biomass was dispensed, transferred to sterile ampoules containing glycerol and frozen (hereinafter referred to as frozen vegetative mycelium (FVM)). This mycelia with a final glycerol concentration of 10-15% was maintained at -75 ° C. Secondary frozen vegetative mycelia (FVM) was prepared by transferring 1.0 milliliter of thawed primary frozen vegetative mycelium (FVM) to inoculum medium 2, incubating for 7 days at 25 ° C and 220 rpm, followed by freezing as described above.

9999

9

999

99

9 9 9 9

9 9 9 9 · 9 * 9 9 9

9 9 9 9

99 9 ·

99 ♦ 9 «

9 9

9 9

9 «9»

2. Inoculum:

After thawing and warming to room temperature, the contents of the frozen ampoule (FVM) MF6239 were used to inoculate the seed cultures at 1.0 milliliter per 50 milliliters of inoculum medium 2. The resulting mixture was grown on a swinging cone shaker at 220 rpm for 7 days. temperature of 25 ° C and relative humidity of 85%.

3. Production:

On solid production medium

An aliquot (10-12 milliliters) of the inoculum was added to 220 milliliters of the liquid portion of the solid production medium 1. To disperse the biomass, the flask was shaken vigorously. The contents of each flask were poured into a 2 liter cylindrical culture vessel containing 675 milliliters of coarse-grained vermiculite. The contents of the culture vessel were shaken / mixed to ensure homogeneous inoculation and homogeneous coverage of the vermiculite. The culture vessel was incubated for 17 days in a horizontal position at approximately 4 rpm on a Wheaton instrument at 22 ° C and 70% relative humidity.

In liquid production medium

Inoculation of the inoculum cultures was performed as described above. Inoculation of 250 milliliters of production medium 1, (1.5 milliliters) of inoculum, of each production flask with a volume of 50 milliliters of liquid was performed in an aliquot. ° C and relative humidity 50-85%.

00 00 ·· * · 00 0 »0 * 00 0 ♦» 0 0 0 0 • 0 · · «0 0 00« * · «

0 000 000 000000 0 # 0 00 0 0000 00 00 00 000 04 00

Sordarin production by fermentation MF6232 (ATCC 74387)

1. Culture:

An oblique section of agar containing MF6232 was aseptically transferred to inoculum medium 1 (50 milliliters in an uncovered flask of volume

250 milliliters). To obtain biomass, the mixture was incubated on a swinging cone shaker for 3 days at 220 rpm, 25 ° C and 85% relative humidity. The biomass was dispensed, transferred to sterile ampoules containing glycerol and frozen (hereinafter referred to as frozen vegetative mycelium (FVM)). These mycelia with a final glycerol concentration of 10-15% were maintained at -75 ° C. Secondary frozen vegetative mycelia (FVM) was prepared by transferring 1.0 milliliter of thawed primary frozen vegetative mycelium (FVM) to inoculum medium 2, the composition of which is shown below, by incubation for 7 days at 25 ° C and 220 rpm and followed by freezing as described above.

2. Inoculum:

The contents of the frozen ampoule (FVM) MF6232, after thawing and warming to room temperature, were used to inoculate the seed cultures at 1.0 milliliter per 50 milliliters of inoculum medium 2. The resulting mixture was grown on a swinging cone shaker at 220 rpm for 7 days. temperature of 25 ° C and relative humidity of 85%.

3. Production:

On solid production medium

An aliquot (10-12 milliliters) of the inoculum was added to 220 milliliters of the liquid portion of solid production medium 2. For fe *

fe fe • • · fe · fefe fefe fefe fefefefe fefe fefe • fefefe fefefe • fefe fefefe fe fefe fe fe * · ·· · fefefe 4 fe · • ·· · ···· • fe »···· ·. · ·· To disperse the biomass, the flask was shaken vigorously. The contents of each flask were poured into a 2 liter cylindrical culture vessel containing 675 milliliters of coarse-grained vermiculite. The contents of the culture vessel were shaken / mixed to ensure homogeneous inoculation and homogeneous coverage of the vermiculite. The culture vessel was incubated for 17 days in a horizontal position at approximately 4 rpm on a Wheaton instrument at 22 ° C and 70% relative humidity.

In liquid production medium

Inoculation of the inoculum cultures was performed as described above. Inoculation of 250 milliliters of production medium 2, (1.5 milliliters) of inoculum.

Each production flask containing 50 milliliters of liquid was made in an aliquot. The flasks were incubated for 7-21 days in a swinging conical shaker at 220 rpm, 25 ° C and 50-85% relative humidity.

Large scale production of sordarin MF6232 (ATCC 74387)

The liquid part of the solid production medium 1 was used in both the inoculum and the production fermenter. After sterilization, cerelose was added to both fermenters at a rate of 30 grams / liter for the seed fermenter and 150 grams / liter for the production fermenter. The inoculum fermentors were inoculated with 2 liters of culture grown in shake flasks. These fermentors were then allowed to grow at 25 ° C for 30 hours until the oxygen uptake was approximately 3 millimoles / liter per hour. After 30 hours, 25 liters of inoculum were transferred to a production fermentor.

reached po • ft ftft • ftft · • · »· • ftftftft ♦ · · • ft ftft • ft ftftftft • ftft • ftftftft ft ft ft ftft · • ft ftftft ·· ftft

I ftft · »ftft ♦ ft ftft ·» ftftft ftft ftft hours at the end

An increase in oxygen uptake of 8-10 millimoles / liter per hour in the production fermentor and cultivation decreased this absorption to 5-7 millimoles / liter per hour. Dissolution of oxygen was performed by increased stirring of the mixture. The pH of the broth was not regulated and generally decreased to 5.3 after 200 hours. The process was performed at 25 ° C.

After 280 hours of growth, the fermentation was terminated and the preparation of the isolation was started. The pH was adjusted to 12 with sodium hydroxide and the mixture was allowed to stand at the fermentation temperature for 20 hours. The pH was then adjusted to 6.0 with sulfuric acid before transferring the mixture to rollers for further processing.

Isolation of sordarin

Insulation I

The methyl ethyl ketone extract from MF6232 culture fermentation (ATCC 74387) corresponding to 64 milliliters of crude broth was evaporated to dryness in vacuo. The obtained 365 milligram residue was dissolved in methanol / dichloromethane (2:98) to a volume of 4.6 milliliters. From this solution, 4.3 milliliters, i.e., a portion containing 341 milligrams of dissolved residue, was collected, and this portion was applied to a flash chromatography column packed with 60 milliliters of silica gel 60 (0.040-0.0630 millimeters, 230-400 mesh, E. Merck) and equilibrated with a 2% solution of methanol in dichloromethane. The column was eluted by step gradient elution and washed successively with 2, 5, 10 and 30% methanol in dichloromethane, using 4 44 4 44 4 44 44 each.

4444 444 40 * 4 • 04 4 0 0 440 4 4 0 0

040040 0 000 14 »

0 00 0 0000 00 00 04 044 0 · 04

240 milliliters of the appropriate solution, and finally the column was washed with 120 milliliters of methanol alone. For each system, 16 fractions of 15 milliliters were collected. The biological test determined that fractions number 39-56 contained a large proportion of the product.

The combined fractions were evaporated to dryness in vacuo to give 103.1 milligrams of the sample, from which a 34.4 milligram portion was taken, which was further purified by HPLC (a 9.4 x 250 millimeter column packed with solid phase). Zorbax Rx-C8, 5 micrometers, which was eluted with a mobile phase consisting of 20% acetonitrile and 80% 0.01 molar aqueous solution of K ₂ HPO ₄ , the pH of which was adjusted to 6.9 with concentrated H ₃ PO ₄ . was set at 4 milliliters per minute at 40 [deg.] C. Detection was performed by diode array). 4 milliliter fractions were collected. Fractions 16-20, which contained a large proportion of product, were combined and concentrated in vacuo to approximately twenty-five percent of the original volume. The concentrate was extracted twice with an equal volume of ethyl acetate, and the organic layers were combined, washed with an equal volume of brine, dried over anhydrous sodium sulfate, and the solvents were evaporated in vacuo to give 3.7 milligrams of sordarin.

Insulation II

The methyl ethyl ketone extract from batch -004Y fermentation of MF6232 culture (ATCC 74387) corresponding to 980 milliliters of crude broth was evaporated to dryness in vacuo. The obtained residue weighing 4.9 grams was dissolved in such an amount of a methanol / dichloromethane mixture (1: 9) that the volume of the solution was 21.5 milliliters. From this solution, 21 milliliters, i.e. a portion containing 4.8 grams of dissolved residue, was taken, and this

0 »0000 • 0 • 00»

0 0 0

0 0 0

0 0 0

0 »00»

0 0

00

0 0 0 00

000

0 »• 0 0 ·

0 0 0

0 0 0

0 0 0

The 0.0000 aliquot was applied to a chromatographic column packed with 500 milliliters of silica gel 60 (0.040-0.0630 millimeters, 230400 mesh, E. Merck) and equilibrated with a 2 percent solution of methanol in dichloromethane. The column was eluted at a rate of 25 milliliters per minute by step gradient elution first with 1 liter of a 2 percent solution of methanol in dichloromethane, then with 1 liter of 5 percent methanol and then with 2 liters of 15 percent methanol. Finally, the column was washed first with 1 liter of 30 percent methanol and then with 2 liters of methanol alone. Twenty-five milliliters fractions were collected. The biological assay determined that fractions 75-85 and 111-121 contained a large proportion of the product and contained compound (I) according to the analysis performed by reverse phase HPLC.

The combined fractions 75-85 and 111-121 were separately evaporated to dryness in vacuo to give samples weighing 69.3 milligrams and 95.3 milligrams, respectively. From the sample obtained by evaporation of the combined fractions 75-85, two 34 milligram aliquots were taken and further purified by two identical HPLC (a 21.2 x 250 millimeter column packed with a 7 micron Zorbax Rx-C8 solid phase was used). was eluted with a mobile phase consisting of 40% acetonitrile and 60% water with a total content of 0.1% H 3 PO 4. The washing rate was set at 20 milliliters per minute at 25 ° C. Detection was performed with a detector at a wavelength of 220 nanometers). 10 milliliter fractions were collected. Fractions 27-31 from the two preparations, which contained a large proportion of product, were combined and concentrated in vacuo to approximately forty percent of the original volume. The concentrate was extracted twice with an equal volume of ethyl acetate and the organic layers were combined, washed with an equal volume of brine, dried over anhydrous sulfate • φ ···· • φ φ

φφφ φ φ φφφ φ · φ φ φ φ φ φ φ · φ φ sodium and after evaporation of the solvents in vacuo, 27 milligrams of sordarin were obtained. Under the same conditions, two 46 milligram aliquots were purified from the sample obtained by evaporation of the combined fractions 111-121. Fractions 25-28 from both preparations were combined and 17 mg of sordarin were obtained after treatment as described above.

Intermediate 1 [IR- (1a, 3aP, 4β, 4a3,7β, 7aa, 83β)] - Benzyl-4-formyl-8a (hydroxymethyl) -4,4a, 5,6,7,7a, 8,8a-octahydro -7-methyl-3- (1-methylethyl) -1,4-methano-s-indacene-3a (1H) -carboxylate (sordaricin benzyl ester)

CHO

To 2 milligrams of sordarin dissolved in 1 milliliter of acetone was added 0.2 milliliter of concentrated hydrochloric acid. The mixture was stirred at room temperature for one day. The solution was diluted with water, treated with aqueous (dichloromethane) and the organic portion was dried over anhydrous sodium sulfate, filtered and concentrated in vacuo. The resulting mixture was dissolved in 2 milliliters of N, N-dimethylformamide (DMF), and 0.1 milliliter of benzyl bromide and excess solid NaHCO 3 were gradually added to the resulting solution. The mixture was stirred overnight at room temperature and then concentrated in vacuo. Chloroform was added to the mixture, and NaHCO ₃ was filtered off. The filtrate was concentrated in vacuo and purified by preparative thin layer chromatography (PTLC) to give 1.0 mg of sordaricin benzyl ester. ¹ H NMR (CDCl ₃ ): δ (1H, m), 3.48 (1H, d (1H, d, J = 11.7), 5.31

): δ 0.51 (3H, d, 1.0 (3H, d, J = 6.6), 2.24 (1 H, m) , 2.51 ·, D, J = 11, 0) , 5.11 04 (1H, d, J = 2.1,)

7.31-7.40 (5 H, m), 9.62 (1 H, s).

Intermediate 2 [IR- (1a, 3aP, 4β, 4a3,7β, 7aa, 83β)] - 4-Methoxybenzyl-4-formyl-8a (hydroxymethyl) -4,4a, 5,6,7,7a, 8,8a -octahydro-7-methyl-3- (1-methylethyl) -1,4-methano-s-indacene-3a (1H) -carboxylate (sordaricin p-methoxybenzyl ester)

The procedure was the same as for the preparation of sordaricin benzyl ester, except that 4-methoxybenzyl chloride was used instead of benzyl bromide. ¹ H NMR (CDCl ₃ ): δ 0.51 (3H, d, · »· fefe fe ·· * • · fe fefe

(1H, s).

Intermediate 3 [IR- (1α, 3a3,4β, 4a3,7β, 7aa, 83β)] - Allyl-4-formyl-8a (hydroxymethyl) -4,4a, 5,6,7,7a, 8,8a-octahydro -7-methyl ^k -3- (1-methylethyl) -1,4-methano-s-indacene-3a (1H) -carboxylate (sordaricin allyl ester)

CHO

The procedure was similar to that for the preparation of sordaricin benzyl ester, except that allyl bromide was used instead of benzyl bromide.

Intermediate 4

Acid [IR- (1α, 3a3,4β, 43β, 7β, 7aa, 8θβ)] - 4-formyl-8a- (hydroxymethyl) -4,4a, 5,6,7,7a, 8,8a-octahydro-7 -methyl-3- (1-methylethyl) -1,4-methano-s-indacene-3a (1H) -carboxylic acid (sordaricin)

CkOH £ H0 xt-rFW

To a methanolic solution of 0.6 milligrams of sordaricin benzyl ester was added Pearlman's catalyst. The mixture was stirred under hydrogen pressure for 15 minutes (pressure created by connecting a lounge which was filled with hydrogen). After filtration through cotton wool and concentration in vacuo, 0.4 milligrams of sordaricin was obtained. 1 H NMR (CDCl ₃ ): δ 0.82 (3H, d, J = 6.8), 0.98 (3H, d, J = 6.6),

1.01 (3H, d, J = 6.9), 1.23 (1H, m), 1.25 (1H, d, J = 12.6), 1.582.10 (9H, m), 2, 34 (1H, m), 2.41 (1H, t, J = 3.6), 3.45 (1H, d, J = 11.0), 4.14 (1H, d, J = 11.0 ), 6.05 (1H, d, J = 3.0), 9.75 (1H,

s).

Intermediate 5 [IR- (1α, 3a8,4β, 4ap, 7β, 7aa, 83β)] - Benzyl-4-cyano-8a (hydroxymethyl) -4,4a, 5,6,7,7a, 8,8a-octahydro -7-methyl-3- (1-methylethyl) -1,4-methano-s-indacene-3a (1H) -carboxylate .0

CN

00

RSS 0 0 0 • · · ·

0 000

0 ·

00

0000 00 00 0 0 0 «0 0 0 · 000 0 0 0 0

0 00 * 00 0 0 0 0 0 0 4 0

400 00 00

161.2 milligrams of sordaricin benzyl ester was dissolved in 6 milliliters of N, N-dimethylformamide, and 1 milliliter of p-methoxybenzyl chloride and excess sodium hydride (50 milligrams of 60% dispersion in mineral oil) were gradually added to the solution. The resulting mixture was stirred overnight, diluted with ether and washed carefully with water. The ether layer was dried over anhydrous sodium sulfate and the volatiles were removed in vacuo. The residue was purified by silica gel chromatography to give 192.5 milligrams (93%) of p-methoxybenzyl ether.

150 milligrams of the obtained ether was dissolved in 5 milliliters of dry ethanol, and 3 milliliters of dry pyridine was added to the solution. After adding 96 milligrams of hydroxylamine hydrochloride, the mixture was heated at 70 ° C for 3 hours. The reaction mixture was cooled and concentrated in vacuo. The residue was dissolved in ether, washed with water and dried over anhydrous sodium sulfate. The residue obtained after removal of the ether in vacuo was purified by preparative thin layer chromatography (PTLC) to give 143.4 milligrams (93%) of the corresponding aldoxime.

143 milligrams of the obtained aldoxime was dissolved in

Excess salt (methoxycarbonylsulfamoyl) was added to the solution and stirred for 2 hours at milliliters of toluene and (700 milligrams) of internal triethylammonium hydroxide.

The mixture temperature is 70 ° C. After concentration in vacuo, the residue was purified by preparative thin layer chromatography (PTLC) to give 116.6 milligrams (84%) of the appropriate nitrile.

67.5 milligrams of the nitrile prepared above was dissolved in 5 milliliters of dichloromethane and 43 milligrams of DDQ and 0.5 milliliter of water were added to the solution. The resulting mixture was stirred at room temperature for 2 hours. After aqueous treatment and ·· ·· ·· ···· ·· ·· ···· ··· ···· • · · · · · ··· · · · · · ··· ·· · Purification by preparative thin layer chromatography (PTLC) afforded 47.6 milligrams (91%) of Intermediate 5: ··· · · · ·· ··· ····· ·· ·· ·· ··· η · ·· .

The following examples are provided to better illustrate the present invention and do not limit the scope of the invention in any way.

Examples of embodiments of the invention

Example 1

Part A equivalent of a compound of the above formula except that R is -CH (C ₆ H ₅ ) 2 and X is -CHO and the preparation of which is described in WO 96/14326 was dissolved in ethanol and the same volume of pyridine and 10 equivalents of hydroxylamine hydrochloride. The reaction mixture was stirred under nitrogen for 20 minutes or until the reaction occurred as determined by analytical thin layer chromatography (TLC), concentrated under reduced pressure and partitioned between water and dichloromethane. The organic portion was dried over anhydrous sodium sulfate, filtered and concentrated in vacuo. The product was obtained after • ·

• ·· ·· ·· purification of the residue by preparative thin layer chromatography (PTLC) in the system (hexane / ethyl acetate).

Part B

To a solution of 1 equivalent of the product obtained in Part A in toluene was added 5 equivalents of Burgess salt. The reaction mixture was stirred at 60 ° C under nitrogen for 1 hour or until sufficient starting material had reacted. Subsequently, the mixture was cooled and the volatiles were removed in vacuo. The product was obtained after purification of the residue by preparative thin layer chromatography (PTLC) in the system (hexane / ethyl acetate).

Part C

The product obtained in part B was dissolved at 0 ° C in dichloromethane containing 2% by weight of trifluoroacetic acid. The resulting mixture was stirred for 4 hours or until sufficient reaction occurred. The volatiles were removed under reduced pressure and the residue was purified by preparative thin layer chromatography (PTLC) to give the desired product of the above formula.

Example 2

[IR- (1α, 3α, 4β, 4β, 7β, 7α, 83β)] - 8α - [(6-deoxy-4-N-methyl-4-D-altropyranosyloxy) methyl] -4-cyano-4 , 4a, 5,6,7,7a, 8,8aoctahydro-7-methyl-3- (1-methylethyl) -1,4-methano-s-indacene3a (1H) -carboxylic (4-cyano-4-deformylsordarin) »· 0 · · • ·

To a solution of 50 milligrams of sordarin in 3 milliliters of N, N-dimethylformamide were added sequentially 0.3 milliliters of benzyl bromide and 200 milligrams of sodium hydride (60% dispersion in mineral oil). The mixture was stirred overnight at room temperature. After aqueous workup (diethyl ether) and purification by preparative thin layer chromatography (PTLC), 2 ', 3-di-C-benzylsordarin benzyl ester was obtained.

To a solution of 1 equivalent of 2 ', 3'-di-Obenzylsordarin benzyl ester, prepared as described above, in ethanol / pyridine (2: 1) was added excess hydroxylamine hydrochlorides and the mixture was stirred at 70 ° C for 2 hours. Subsequently, the mixture was concentrated in vacuo and after aqueous workup (dichloromethane) and subsequent purification by preparative thin layer chromatography (PTLC), 2 ', 3'-di-O-benzyl-4-aldoximsordarin benzyl ester was obtained.

To a solution of 2 ', 3'-di-O-benzyl-4-aldoximsordarin benzyl ester, prepared as described above, in toluene was added an excess of (methoxycarbonylsulfamoyl) triethylammonium hydroxide inner salt (Burgess reagents). The resulting mixture was stirred at 70 ° C under nitrogen for 2 hours. Concentration in vacuo and purification by preparative thin layer chromatography (PTLC) gave 2 ', 3'-di-O-benzyl-4-cyano-4-deformylsordarin benzyl ester.

·· ·· ·· ···· ·· 00 • 00 0 · 0 0000 • 0 · · 0000 0 0 0 0

60 0 0000 00 0 000 00 · 00 0 00 0 0000 00 00 00 000 00 00 A solution was prepared above prepared benzyl ester in methanol. The solution was added palladium hydroxide on carbon (Pearlman 's catalyst) a vessel was flushed with hydrogen.

The mixture was stirred vigorously under a hydrogen atmosphere for 15 minutes, filtered and concentrated to give the product from the beginning of this example. MS (CI): m / z = 507.5 (M + NH ₄ ).

Example 3

[IR- (1α, 3a3,4p, 4ap, 7p, 7aa, 8ββ)] -8α - [(6-deoxy-4-D-altropyranosyloxy) methyl] -4-cyano-4,4a, 5,6,7 , 7α, 8,8α-octahydro-7-methyl-3- (1-methylethyl) -1,4-methano-s-indacene-3a (1H) carboxylic (4-cyano-4-deformyl-4'-demethyl-sordarin)

One milliliter of frozen Streptomyces avermitilis MA 4848 mycelia (ATCC 31272) was inoculated into each of eight capped Erlenmeyer flasks containing 40 milliliters of BaSa medium, [which contained 20 grams of Difco extract, 20 grams of Hykasa (salt-free), Sheffield ), 20 grams of dextrose, 2 grams of potassium nitrate, 10 milliliters of a trace element mixture as defined below, which has a pH of 7.0 and which has been heat-treated for 20 minutes • · • · ···· · · ·· • · · ··· · · «· · • · · · ···· · · · f • ······ · ··· · · · • · · ·· · ···· • · · · ·· · · · ·· · · in an autoclave]. The flasks were incubated at 27 ° C at 220 rpm microscopically for non-mycelial origin.

hours at temperature and pH 6.7-6.8. Mycelia presence of impurities

Component

Content in 1 liter

NaCl (12.5% solution) 4 ml

MgSO ₄ .7 H ₂ O (12.5% solution) 4 ml

FeSO ₄ .7 H ₂ O 25 mg

MnSO ₄ .H ₂ O (0.5% solution) 1 ml

ZnSO ₄ .7 H ₂ O (1% solution) 1 ml

CaCl ₂ .2 H ₂ O (2% solution) 1 ml

Forty milligrams of the product prepared in Example 2 (4-cyano-4-deformylsordarin) was dissolved in 0.40 milliliters of 80% ethanol and 50 microliters of this solution was added to each flask. The mixture was incubated at 27 ° C for 18 hours. The course of the reaction and the determination of its end were performed by analytical HPLC. 200 milliliters of broth were diluted with an equal volume of methanol, extracted and the solids removed by centrifugation. Most of the methanol was removed from the supernatant in vacuo. The pH of the remaining approximately 200 milliliter solution was adjusted to 11 with sodium hydroxide. The solution was extracted twice with 200 milliliters of dichloromethane and the pH of the aqueous layer was adjusted to 2.5 with dilute sulfuric acid. The aqueous layer thus treated was then extracted twice with dichloromethane. The combined dichloromethane layers were washed successively with water, brine and dried over anhydrous sodium sulfate. Sodium sulfate was filtered off and dichloromethane was removed in · · · · · · · · · · • · · ··· * · φ · • · · · · · · · fe · fefe · fefefefe ·· fefe fefe fefefe fefe fefe vacuum. 53 milligrams of solid residue was obtained, which was purified by reverse phase preparative HPLC on a Phenomenex Primesphere C8 stationary phase with a particle size of 5 micrometers, which was packed with a column measuring 9.4 x 250 millimeters. The mobile phase was an acetonitrile / water mixture (34:66) containing 0.1% phosphoric acid and the washing rate was 3.5 milliliters per minute at 40 ° C. The product started to flow out of the column after 14.4 minutes. The product-rich fractions were combined and the acetonitrile was removed from the mixture in vacuo under a stream of nitrogen. The remaining aqueous solution was extracted with dichloromethane as described above to give 7.2 mg of the desired product.

^{Χ Η} NMR (1.5 mg in 0.25 mL CD ₃ OD): δ 0.790 (3H, d, J = 6.8), 1.059 (3H, d, J = 6.8), 1.161 (3H, d , J = 6.8), 1.141.32 (m), 1.257 (3H, d, J = 6.4), 1.650 (1H, m), 1.74-1, 86 (m),

2.00-2.18 (m), 2.43-2 (IH, dd, J = 4.4; 12.8), 2,670 (IH, no heptet, J = 6.8), 2,836 th most common (IH, brdd, J = 3.6) , 3,450 (IH, dd , J = 4 , 0; 9.2), 3.67 (2H, m), 3, 871, (1H, dd, J = 3, 6; 3.6) , 3,914 (IH, d, J = 9, 6), 4.472 (brs), 6, 225 (brd, J = 3, 2). ID (thin film on

ZnSe): 2958, 2234, 1713, 1071 cm- ¹ . MS: 475.2607 (M ^& lt ^{; +} & gt ^; )

Example 4

• · • · • »• · • ·

Part A

0.500 grams (3.37 millimoles) of digitoxose was azeotroped with dry benzene in a flask. The thus treated material was dissolved in 5 milliliters of dry pyridine, and 5 milliliters of acetic anhydride was added to the resulting solution. The reaction mixture was stirred for 18 hours. The reaction was monitored by thin layer chromatography on silica gel in ethyl acetate / hexane (1: 2) until complete disappearance of digitose. The volatiles were removed in vacuo to give 1.013 grams of a pale yellow oil. ¹ H NMR (CDCl 3): δ 1.11 (3H, d), 1.96-2.16 (2H, m), 2.02 (3H, s), 2.11 (6H, s), 4, 06 (1H, m), 4.61 (1H, dd), 5.49 (1H, bq), 6.02 (1H, dd). MS: 215.1 (MC ₂ H ₃ O ₂ ) ⁺ .

Part B

9.31 grams (33.9 millimoles) of peracetyldigitose prepared as described in Part A was added to 150 milliliters of water and 50 milliliters of glacial acetic acid was added to the resulting mixture. The reaction mixture was stirred at room temperature for three days. After removal of the solvent, the crude residue was purified by flash chromatography on silica gel eluting with ethyl acetate / hexanes (1: 1). 7.37 g (94%) of a light yellow syrup were obtained, which was identified as a mixture of 3,4-diacetoxydigitose anomers.

Part C

1.23 grams (5.3 millimoles) of 3,4-diacetoxydigitose was weighed into a flask and 25 milliliters of anhydrous dichloromethane was added under nitrogen. Then, 0.35 grams (1.1 millimoles) of cesium carbonate and 7.2 grams (50 millimoles) of trichloroacetonitrile were added sequentially to the mixture. The mixture was stirred for about 1 hour, filtered and the volatiles were • 0 · · • ·

0

000 removed in vacuo. 2 grams of crude trichloroacetamidate was isolated and used without purification in further reactions. 1 H NMR (CDCl ₃ ): δ 1.34 (3H, d), 2.03 (3H, s), 2.07 (3H, s), 2.11 (1H, m), 2.29 (1H, ddd), 4.17 (1H, m), 4.80 (1H, dd), 5.52 (1H, m), 6.20 (1H, dd), 8.76 (1H, s).

Part D

1.6 grams (3.8 mmol) of intermediate 1 was azeotroped with benzene and the dried compound was dissolved in 10 ml of dichloromethane. 0.340 grams (1.5 millimoles) of anhydrous zinc bromide was added to the solution. The mixture was cooled to 0 ° C and added via a linear syringe dispenser over one hour. 2.8 grams (7.5 millimoles) of Part C product dissolved in 5 milliliters of dichloromethane. The reaction mixture was then poured into aqueous sodium bicarbonate solution and extracted with dichloromethane. The organic layer was dried over anhydrous sodium sulfate, filtered and the solvents were removed in vacuo. The crude product was purified by flash chromatography on silica gel eluting with ethyl acetate / hexane (1: 4) to give 2.09 grams (86%) of the desired product as a clear oil. The product was determined to be a 2: 3 mixture of aa β anomer. MS: 654.2 (M + NH ₄ ) ⁺ .

Part E

2.09 grams (3.3 millimoles) of the product from Part D was dissolved in 50 milliliters of methanol and 0.20 grams of potassium carbonate was added to the solution. The reaction mixture was stirred at ambient temperature for 2 hours. Potassium carbonate was filtered off and the volatiles were removed in vacuo. The crude product was

00 00 0000 00 00 0000 000 00 * 0 0 00 0 0 0000 0 00 0 0 0 000 00 0 000 00 0 00 0 0 0 0 0000 00 · 0 00 000 00 00 purified by flash chromatography on silica gel eluting first with ethyl acetate / hexane (1: 2), later ethyl acetate / hexane (1: 1) to give 0.84 grams (49%) of the desired deacetylated β-anomer along with 0.48 grams (26%) of the desired deacetylated α-anomer.

α-Anomer: partial ^X H NMR (CDCl ₃ ): δ 0.51 (3 H, d), 0.80 (3H, d), 0.85-0.95 (m), 1.00 (3H, d), 1, 02 (m), 1.14 (3H, d), - 2.77 (1H, t), 3.44 (1 H, d), 3.58 (m), 3.93 (1H, dd), 3.27 a 3, 98 (2H, TIB Quartet), 4.7 5 (1H, m), δ , 18 (2H, AB quartet), 6, 02 (1H, m), 7.37 (5H, m), 9, 67 (1 H, s). MS: 570.5 (M + NH ₄ ) ⁺ β-Anomer: partial ¹ H NMR (CDCl 3): δ 0.51 (3 H, d), 0.81 (3H, d), 0.85-1.1 (m), 1.00 (3H, d), 1.13 (3 H, d), 2.83 (1H, t), 3.64 (1H, m), 3.60 a 3.87 (2H, TIB Quartet), 4.10 (1H, bs), 4.58 (1H, dd), 5.17 (2H, AB quartet), 6.01 (1H, d), 7.35 (5H, m,), 9.69 (1 H, s). MS: 423.3 (M + H-digitose)

Part F

35.6 milligrams of the β-anomer prepared in Part E was dissolved in 3 milliliters of dibromoethane, and 3 milliliters of 50% aqueous sodium hydroxide solution and 4.2 milligrams (0.013 millimoles) of tetrabutylammonium bromide were added sequentially to this solution. The reaction mixture was stirred vigorously for 18 hours, extracted with dichloromethane and the organic layer was dried over anhydrous sodium sulfate, filtered and the volatiles removed in vacuo. The residue was purified by preparative thin layer chromatography (PTLC) in hexane / ethyl acetate (9: 1). 16.4 milligrams (45%) of a white solid were obtained. Partial ^X 1 HNMR (CDCl _3): δ 0.52 (3H, d), 0.90 (3H, d), 0.9 (m), 1.00 (3H, d), 1.04 (m) , 1.13 (3H, d),

66 • ♦ 9 9 • 9 9 • 9 9 • 9 99 • · 99 99 9 9 9999 9 9 9 9 9 9 9999 9 9 9 9 9 9 9 9 9 9 9 9 9 99 9 9 9 9 • 9 9 · 9 9 9 9 9 9 9 9 9 9 9 9 99 »9 2.71 (IH, t), 3.38 (1H, m), 3, 60 a 3, 89 (2H, AB quartet), 3.64 (IH, m), 4.13 (IH, m), 4.48 (IH, dd) , 4.85 (1H, s), 5.14 (IH, s), 5.18 (2H, AB quartet), 6.01 (IH , d), 7.36 (5H, m), 9, 71

(1H, s).

Part G

9.7 milligrams (0.017 millimoles) of the product prepared in Part F was dissolved in 1 milliliter of pyridine, and to this solution were added successively 1 milliliter of ethanol and 12 milligrams (0.17 millimoles) of hydroxylamine hydrochloride. The reaction mixture was heated to 70 ° C and stirred for 1 hour. The mixture was cooled and the solvent was removed in vacuo. The residue was partitioned between water and dichloromethane, and the aqueous layer was further extracted with dichloromethane. The combined organic layers were dried over anhydrous sodium sulfate, filtered and the volatiles were removed in vacuo. Purification by preparative thin layer chromatography (PTLC) in hexane / ethyl acetate (4: 1) gave 8.6 mg (87%) of solid product. Partial

^{Χ Η} NMR (CDC1 ₃₎ : δ 0 , 52 (3H, d), 0 , 81 * 3H, d), 0.9 (m), 0.98 (3H, d), 1.01 (m), 1.23 (3 H, d), 1.34 (1H, d), 2.62 (1H, t), 3.36 (1H, m), 3.60 (1H, d), 3.64 (IH, dd), 3.87 (1H, d), 4.13 (1H, m), 4.49 (IH, dd), 4.85 (1H, s,), 5.15 (1H, s), 5.18. (2H, AB quartet), 5, 96 (1H, d), 7.36 (5H, m), 7.46 (1H, bs), 7.81

(IH, s)

Part H

8.6 milligrams (0.015 mmol) of the product prepared in Part G was dissolved in 2 milliliters of toluene and 18 milligrams (0.074 mmol) of Burgess reagent was added to this solution. The reaction mixture was stirred at 60 ° C for about 1 hour. Additional Burgess reagent was added to the mixture and the mixture • · 999 · «* · • · 0 · · ♦ · · ··» * · <

»· · 4 ·· ·· was stirred for another 20 minutes, cooled and the solvent was removed in vacuo. Purification of the residue by preparative thin layer chromatography (PTLC) in hexane / ethyl acetate (4: 1) gave 7.3 milligrams (86%) of solid product. Partial ¹ H NMR (CDCl 3): δ 0.39 (3H, d), 0.88 (3H, d), 0.95 (m), 1.14 (3H, d), 1.23 (3H, d) ), 2.62 (1H, m), 2.72 (1H, t), 3.36 (1H, m), 3.59 (1H, d), 3.63 (1H, dd), 3.88 (1H, d), 4.11 (1H, m), 4.48 (1H, dd), 4.85 (1H, s), 5.14 (1H, s), 5.18 (2H, AB quartet ), 6.12 (1H, m), 7.34 (3H,

m), 7.46 (2H, m).

Part I

7.3 milligrams (0.013 mmol) of the product prepared in Part H was dissolved in 2 milliliters of methanol, and 2 milligrams of Pearlman's catalyst was added to the solution. The reaction vessel was purged with hydrogen and stirred under a hydrogen atmosphere for 2 hours. The mixture was filtered through celite and the volatiles were removed in vacuo to give 6.1 mg (100%) of the final product as a white solid. Partial ¹ H NMR (CDCl ₃ ): δ 0.77 (3H, d), 1.01 (3H, d),

1, 18 (3H, d), 1.28 (3H, d), 2.23 (1H, dt), 2.35 (1H, dd), 2.67 (2H, m), 3.39 (1H, m), 3.54 (1H, d), 3.63 (1H, dd), 3.99 (1 H, m), 4.15 (1H, m), 4.57 (1H, dd), 4.85 (1H r s), 5.13 (1H, s),

6.16 (1 H, d). MS: 489.2 (M + NH ₄ ) ⁺ . IR: 2240 cm- ¹ ,

φ φ φ «φ φ φ φ φφ

ΦΦΦ φφ φφ • · φ φ • φ φ φ φ φφφφ • φ φ φφ • φ φ φ φ φ

Example 5

Part A

0.84 grams (1.5 mmol) of the β-anomer prepared in Part E of Example 4 was dissolved in 100 milliliters of toluene, 0.568 grams (2.3 mmol) of dibutyltin oxide was added to the solution under nitrogen, and the mixture was heated to reflux for 4 hours. and mixed. Then, the reaction mixture was cooled to room temperature, and 0.544 grams (4.5 mmol) of allyl bromide and 2.3 milliliters of a 1.0 molar solution of tetrabutylammonium fluoride in tetrahydrofuran (2.3 mmol) were added sequentially. The mixture was heated to 50 ° C, cooled after 36 hours and the volatiles removed in vacuo. The residue was purified by flash chromatography eluting with a gradient of hexane / ethyl acetate (9: 1, 4: 1, 2: 1) to give 0.454 (51%) of UV absorbing product.

Partial NMR (CDCl ₃ ) : δ 0.51 (3H, d), 0.81 (3H, d), 0.9 (m), 1.00 (3H, d), 1.05 (m), 1.22 (3H, d), 2.21 (1 H, m), 2.72 (1H, t), 3.01 (1 H, dd), 3.60 (1 H, d), 3.68 (1 H, m), 3.87 (1H, d), 3.98 (1H, m), 4.10 (1 H, m), 4.18 (1H, m), 4.59 (1H, dd), 5.10 (m), 5.18 (2H, AB quartet), 5.26 (1H, d), 5.86 (1H, m), 6.01 (1H, d), 7.37 (5H, m), 9.66 ( 1H, S) •

* * 9 49 4 (0.49 mmol) of the product prepared ♦ ♦ 99 ♦ 9 9 4

9 4 4

4 4 44

4 4 · ί

4

444

44 »*» «> 4 4 I

I ♦ · 1 ► · · 4 · «·«

Part Β

A mixture of 288 milligrams in Part A, 511 milligrams (1.95 mmol) of triphenylphosphine and

133 milligrams (1.95 mmol) of imidazole was dissolved in 40 milliliters of freshly distilled tetrahydrofuran in a flask. To the solution was added 371 milligrams (1.46 mmol) of solid iodine, and the mixture was stirred under a nitrogen atmosphere for 1.5 hours. Then, 1 N hydrochloric acid was added to the mixture, and the reaction mixture was extracted with ethyl acetate. The organic portion was washed successively with water, sodium thiosulfate solution and brine. Subsequently, the organic portion was dried over anhydrous sulfate, filtered and concentrated in vacuo. The residue was prepared by preparative thin layer chromatography (PTLC) in hexane / ethyl acetate (9: 1) to give 211 mg (61%) of sodium, purified

product. Partial ¹ H NMR (CDCl ₃ ): δ 0.51 (3H, d), 0.80 (3H, d) 0.9 (m), 1.00 (3H, d), 1.05 (m), 1.33 (3H, - d), 1.22 (1H, d) 2.51 (1 H, dd), 2.70 (1 H, t), 3.13 ! (1H, t), 3.25 (1H, m), 3.59 and 3.82 (2H, AB quartet), 4.01 (1H, r m), 4.38 (1H, dd), 5.18 (2 H, AB quartet), 5.20 (1 H, s), 5.30 (1H, d), 5.96 (1H, m), 6.00 (1H, d), 7.37 (5H, m), 9.69 (1H, s). MS: 725.0

(M + Na).

Part C

To 8 milliliters of dry toluene was added 0.267 milliliters (0.993 mmol) of tri-n-butyltin hydride and the solution was heated to reflux for approximately 2 hours. 211 milligrams (0.301 mmol) of the product prepared in Part B was dissolved in 10 milliliters of dry toluene and added to the refluxing tri-n-butyltin hydride solution over 2 hours using a linear syringe dispenser. An additional equivalent of tri-n-butyltin hydride was added to the mixture and the mixture was heated further

ΊΟ · 0 «0 0000 00 00 * 0 0 0 000 0000 • · 0 0« «0 0« 0 «0 ·

0 000 00 0 0 «0 0« 0

0 «0 · 0 0 0000

00 «0 000 00 00 minutes, when it was determined by analytical thin layer chromatography (TLC) that all the starting compound had reacted. The mixture was cooled and the volatiles were removed under reduced pressure. Purification by preparative thin layer chromatography (PTLC) gave a yellow oil which was further purified by preparative HPLC (Zorbax Rx-C8 solid phase eluting with 95% acetonitrile and 5% water). wavelength 220 nanometers) and 2 products were obtained, namely 23.0 milligrams (13%) of β-anomer and α-anomer.

β-anomer: Partial ^X 1 HNMR (CDCl _3): δ 0.50 (3H, d), 0.80

(3H, d), 0.89 (m), 0.98 (3H, d), 1.02 (3H, d), 1.20 (3H, d) 2.22 (1 H, m), 2.72 (1H, t), 3.29 (2H, m), 3.58 (1H, d), 3.65 (1H, t), 3.82 (1H, d), 3 , 99 (1H, t), 4 , 41 (1H, dd), 5.19. (2H,

AB quartet), 6.01 (1H, m), 7.36 (5H, m), 9.72 (1H, s).

α-anomer: Partial ^X 1 HNMR (CDCl _3): δ 0.47 (3H, d), 0.79

(3H, d), 0.89 (1H, m), 0.93 (3H, d), 0.98 (3H, d), 1.21 (3H, d), 2.21 (1H, m), 2.38 (1H, m), 2.66 (1H, t), 3.39 (1H, t), 3.54 (1H, d), 3.65 (1H, t), 3.77 (1H, d), 3.82 (1H, t), 4.64 (1H, m), 5.18 (2H, S), 5.99 (1H, m), 7.32 (5H, m)., 9, 69 (1H,

s).

Part D

5.6 milligrams in part C of the solution was

6.7 milligrams A reaction mixture (0.0097 millimoles) of the β-isomer prepared was dissolved in 0.5 milliliters of ethanol, and 0.5 milliliters of pyridine and (0.097 millimoles) of hydroxylamine hydrochloride were added sequentially. was stirred in the atmosphere for about 20 minutes

Μ 99 • · · · • 9 · 9 • · ··· 9

9 9 «9 99

9

9

9 ·· · Μ · • 9 • 999

9 «99

9 9 9 * 9 9 9

9 9 »9

9 9 9

99 nitrogen, concentrated under reduced pressure and partitioned between water and dichloromethane. The organic phase was dried over anhydrous sodium sulfate, filtered and concentrated in vacuo. Purification of the residue by preparative thin layer chromatography (PTLC) in hexane / ethyl acetate (4: 1) gave 6.6 milligrams (quantitative yield) of the desired product. Partial

¹ H NMR (CDCl 3) : δ 0, 51 ( 3H, d), 0, 81 (3H, d), 0.98 (3H, d), 1.02 (3 H, d), 1.20 (3H, d), 2.23 (1H, m), 2.63 (1H, t), 3.29 (2H, m), 3.58 (1H, d), 3.65 (1 H, t), 3.82 (1H, d), 3, 99 (1H, t), 4.41 (1H, dd), 5.19 (2H, s), 5.95 (1H , m), 7.35 (5H, m), 7.47 (1 H, s); 7.81 ( 1H, s).

Part E

To a solution of 6.6 milligrams (0.011 millimoles) of the product prepared in Part D in 1 milliliter of toluene was added 13.3 milligrams (0.055 millimoles) of Burgess salt. The mixture was stirred at 60 ° C under nitrogen for one hour, cooled and the volatiles removed in vacuo. Purification of the residue by preparative thin layer chromatography (PTLC) in hexane / ethyl acetate (9: 1) gave 5.1 milligrams (81%) of the desired product. Partial ’NMR (CDCl 3) · δ

0.39 (3H, d) 0, 86 (3H, d), 1.01 (3H, d), 1.15 (3H, d), 1.20 (3H, d), 2.27 (1H, dd), 2, 63 (1H, m), 2.73 (1H, t), 3.29 (1H, m), 3.58 (1 H, d), 3, 65 (1H, t), 3.83 (1H, d), 3.99 (1H, t), 4.41 (1H, dd), 5.27 '(2H, AB quart et), 6, 13 (1 H, m), 7.33 (3H,

m), 7.46 (2H, m).

Part F

To a solution of 5.1 milligrams (0.0087 millimoles) of the product prepared in Part E in 1 milliliter of methanol was added ♦ 0 00 · »···· ► 0 0 0 0 0 0

P 0 0 0 0 0 000 »0 000 000 *

00

000

00 »0 0 0

I 0 0 0

I 0 0 0> 0 0 0

00 approximately 1 milligram of palladium hydroxide on carbon (Pearlman's catalyst). The reaction vessel was purged with hydrogen and the mixture was stirred vigorously under pressure for 30 minutes

101.325 kilopascals (1 atmosphere) in a hydrogen atmosphere. The catalyst was removed by filtration through celite, and after concentrating the filtrate under reduced pressure,

3.8 milligrams (88%) of the final product as a white solid. Partial ^X 1 HNMR (CDCl _3): δ 0.77 (3H, d), 1.01 (3H, d)

1.04 (3H, d), 1.02 (3H, d), 1.17 (3H, d), 1.22 (3H, d), 3.67 (1H, t), 3.98 (1H , t), 4.05 (1H, d), 4.55 (1H, dd), 6.17 (1H, m).

Example 6

Part A

18.3 milligrams (0.033 millimoles) of the α-anomer prepared in Part E of Example 4 was dissolved in 1.5 milliliters of dibromoethane, and 1.5 milliliters of 50% aqueous sodium hydroxide solution (0.006 millimoles) of tetrabutylammonium bromide was gradually added to the resulting solution.

The mixture was stirred vigorously for hours, and after the addition of another 6 milligrams and 2.1 milligrams, the reaction mixture was fefe fefe fefe

The mixture was stirred for a further 24 hours in a mixture of tetrabutylammonium bromide. The reaction was quenched by the addition of 9.4 mL of 2 N hydrochloric acid and partitioned between water and dichloromethane. The combined organic layers were dried over anhydrous sodium sulfate, filtered and concentrated in vacuo. The residue was purified by preparative thin layer chromatography (PTLC) in hexane / ethyl acetate (4: 1) to give 3.1 mg (17%) of product. Partial ^X 1 HNMR (CDCl _3): δ 0.51 (3H, d), 0.81 (3H, d), 0.9 (m), 1.00

(3H, d), 1, 05 (m), 1.13 (3H, d), 2.78 (1H, t), 3.63 (2H, 3.23 and 3.91 (2H, AB quartet), 4.05 (1H, m), 4.62 (IH, m), (IH, s), 5, 13 (IH, s), 5.18 (2H, AB quartet), 5.99 (IH, 7.35 (5H, m) , 9.69 ( IH, s).

Part B

To a solution of 3.1 milligrams (0.0055 millimoles) of the product prepared in Part A in 1 milliliter of methanol was added approximately 1 milligram of palladium hydroxide on carbon (Pearlman's catalyst). The reaction vessel was purged with hydrogen and the mixture was stirred under a hydrogen atmosphere for 40 minutes. The catalyst was removed by filtration through celite, and the filtrate was concentrated under reduced pressure to give 3.1 milligrams (quantitative yield) of the final product. Partial ^X H NMR (CDCl3): δ 0.80 (3H, d), 0.95 (3H, d), 1.01 (3H, d)

1.27 (3H, d), 2.39 (m), 2.78 (1H, t), 3.76 (4H, m), 4.06 (1H, m), 4.71 (1H, m) ), 4.87 (1H, s), 5.13 (1H, s), 6.02 (1H, d), 9.73 (1H, s).

• · «· • · • · • ·« «· · ftft ·· ft ftft · · ftft • ftft · · ···· • · ftftft ftftft ft • · ftftft ft • ft ftft ftft ftftft

Example 7

Part A The equivalent of the α-isomer prepared in Part C of Example 5 was dissolved in ethanol, and an equal volume of pyridine and 10 equivalents of hydroxylamine hydrochloride were added sequentially to this solution. The reaction mixture was stirred under nitrogen for about 20 minutes or until sufficient starting material was shown to be reactive by analytical TLC. The reaction mixture was concentrated under reduced pressure and partitioned between water and dichloromethane. The organic phase was dried over anhydrous sodium sulfate, filtered and concentrated in vacuo. Purification of the residue by preparative thin layer chromatography (PTLC) in hexane / ethyl acetate gave the desired product.

Part B

To a solution of 1 equivalent of the product prepared in Part A in toluene was added 5 equivalents of Burgess salt. The mixture was stirred at 60 ° C for about one hour, or until the starting material had reacted, under a nitrogen atmosphere, cooled and the volatiles removed in vacuo. Purification of the residue by preparative thin layer chromatography (PTLC) in hexane / ethyl acetate gave the product.

·· * ·· · • ft ftft • ftftft · · · · · * · • II · · ftft ··· · ft · · ft ······ · ftftft ft · ft • ft · ftft · · · · ft • ft ftft ftft ftftft ftft ftft

Part C

To a solution of the product prepared in Part B in methanol was added a catalytic amount of palladium hydroxide on carbon (Pearlman's catalyst). The reaction vessel was purged with hydrogen and the mixture was stirred vigorously under pressure

101.325 kilopascals (1 atmosphere) in a hydrogen atmosphere for 30 minutes or until the starting compound was completely reacted. The catalyst was removed by filtration through celite, and the filtrate was concentrated under reduced pressure to give the final product.

Example 8

Part A

16.3 milligrams (0.039 mmol) of intermediate 5 were azeotroped three times with benzene and the dried compound was added to the flask together with 2.0 grams (0.0078 mmol) of anhydrous zinc bromide under nitrogen. Then, 1.0 ml of dry dichloromethane was added to the flask, and the mixture was cooled to 0 ° C. A solution of 29.2 milligrams (0.078 millimoles) of 3,4-diacetyldigitose-1-trichloroacetamidate (prepared by the method) was added to the reaction mixture over one hour.

9 9 9

0 999

9 9

99

9 9

9 999

9 9 0

0 0

0 0 00 • 0 00

0 0 0

0 0 0

0 0 0 0

0 0 0

04 as described in Part C of Example 4) in 0.075 milliliters of dry dichloromethane. The mixture was allowed to warm to room temperature overnight, poured into saturated aqueous sodium bicarbonate solution and extracted with dichloromethane. The organic phase was dried over anhydrous sodium sulfate, filtered and the volatiles were removed in vacuo. The residue was purified by preparative thin layer chromatography (once in hexane / ethyl acetate (9: 1), then in hexane / ethyl acetate (4: 1)) to give 17.0 milligrams (69%) of product as a mixture and - and the β-isomer in a ratio of about 1: 2.

Part B

The product prepared in Part A was dissolved in methanol and to this solution. palliium hydroxide on carbon (Pearlman's catalyst) was added. The reaction vessel was purged with hydrogen and the mixture was stirred vigorously under a hydrogen atmosphere for 30 minutes or until the starting material was completely reacted. The catalyst was removed by filtration through celite, and the filtrate was concentrated under reduced pressure to give the final product. MS: 561.4 (M + NH ₄ ) ⁺ .

Example 9

R = -Η, X = -CN ·· ·· »» ·· 99

9 9 9 9 9

9 9 9 9 9

9 999 9 9 9

9 9 9 9

99 99

99

9 9 9 9

999 · 9 9 9 • 9 9 · 9 · • 9 9 9 ·

999 99 99

Part A

One equivalent of a compound of the above formula wherein R is -CH (C ₆ H ₅ ) ₂ and X is -CHO, the preparation of which is described in published patent application WO 96/14326 was dissolved in ethanol and an equal volume was added gradually to this solution. pyridine and 10 equivalents of hydroxylamine hydrochlorides. The reaction mixture was stirred under nitrogen for about 20 minutes or until sufficient amount of starting material was shown to be reactive by analytical TLC. The reaction mixture was concentrated under reduced pressure and partitioned between water and dichloromethane. The organic phase was dried over anhydrous sodium sulfate, filtered and concentrated in vacuo. Purification of the residue by preparative thin layer chromatography (PTLC) in hexane / ethyl acetate gave the desired product.

Part B

To a solution of 1 equivalent of the product prepared in Part A in toluene was added 5 equivalents of Burgess salt. The mixture was stirred at 60 ° C for about one hour, or until sufficient starting material had reacted, under a nitrogen atmosphere, cooled, and the volatiles removed in vacuo. Purification of the residue by preparative thin layer chromatography (PTLC) in hexane / ethyl acetate gave the product.

Part C

To a solution of the product prepared in Part B in methanol was added a catalytic amount of palladium hydroxide on carbon (Pearlman's catalyst). The reaction vessel was purged with hydrogen and the mixture was stirred vigorously under pressure. · 9 • 9 9 9 • · · · • 9 999 9

9 · »9 ··

9999 »*

9 9 * ·· • C • 9 «· •« «

9 9

9 9 • «9

9 9

101.325 kilopascals (1 atmosphere) in a hydrogen atmosphere for 30 minutes or until the starting compound was completely reacted. The catalyst was removed by filtration through celite, and the filtrate was concentrated under reduced pressure to give the final product.

Example 10

Part A

One equivalent of a compound of the above formula wherein R is -CH (C ₆ H ₅ ) ₂ and X is -CHO, the preparation of which is described in published patent application WO 96/14326 was dissolved in ethanol and the same volume of pyridine and 10 equivalents of hydroxylamine hydrochloride. The reaction mixture was stirred under nitrogen for about 20 minutes or until sufficient amount of starting material was shown to be reactive by analytical TLC. The reaction mixture was concentrated under reduced pressure and partitioned between water and dichloromethane. The organic phase was dried over anhydrous sodium sulfate, filtered and concentrated in vacuo. Purification of the residue by preparative thin layer chromatography (PTLC) in hexane / ethyl acetate gave the desired product.

79 00 00 ·· • · · · · 0 • 0 · 0 0 · • · ··· · · 0 0 0 0 0 0 • 0 ·· ·· • 000 00 • · • 00 0 0 0 0 0 0 ··· · « • 0 • · 0 • 0 0 • 0 0 0 0 0 • 0 Part B To solution 1 product equivalent prepared in parts A v toluene was added 5 equivalents Burgess salts. Mixture was mixed at temperature 60 ° C for about one hour, , or so

until a sufficient amount of the starting compound had reacted, in a nitrogen atmosphere, cooled and the volatiles removed in vacuo. Purification of the residue by preparative thin layer chromatography (PTLC) in hexane / ethyl acetate gave the product.

Part C

The product prepared in Part B was dissolved at 0 ° C in dichloromethane containing 2% by weight of trifluoroacetic acid. The resulting solution was stirred for 4 hours or until the starting compound was completely reacted. The volatiles were removed in vacuo. Purification of the residue by preparative thin layer chromatography (PTLC) gave the final product.

Example 11

Part A

One equivalent of zofimarin (where R = -H and X = -CHO) was dissolved in dry N, N-dimethylformamide (DMF) containing 5% by volume of p-methoxybenzyl bromide. Excess solid sodium bicarbonate was then added to the solution and the mixture was stirred at room temperature for 18 hours or until a substantial amount of zofimarin had reacted. The mixture was concentrated in vacuo, and chloroform was added to the residue. Sodium bicarbonate was removed from the mixture by filtration, the filtrate was concentrated in vacuo, and the residue was purified by preparative thin layer chromatography (PTLC) to give the product of the above formula wherein R is -CH ₂ C ₆ H ₄ -p-OCH ₃ and X is -CHO.

Part B

One equivalent of the product prepared in Part A was dissolved in ethanol, and to this solution was added successively an equal volume of pyridine and 10 equivalents of hydroxylamine hydrochloride. The reaction mixture was stirred under nitrogen for about 20 minutes or until sufficient starting material was shown to be reactive by analytical TLC. The reaction mixture was concentrated under reduced pressure and partitioned between water and dichloromethane. The organic phase was dried over anhydrous sodium sulfate, filtered and concentrated in vacuo. Purification of the residue by preparative thin layer chromatography (PTLC) in hexane / ethyl acetate gave the desired product.

Part C

To a solution of 1 equivalent of the product prepared in Part B in toluene was added 5 equivalents of Burgess salt. The mixture was stirred at 60 ° C under nitrogen for about one hour, or until sufficient starting material had reacted, cooled and the volatiles removed in vacuo. Purification of the residue by preparative thin layer chromatography (PTLC) in hexane / ethyl acetate gave the product.

Part D

The product prepared in Part C was dissolved at 0 ° C in dichloromethane containing 2% by weight of trifluoroacetic acid. The resulting solution was stirred for 4 hours or to completely react the starting material. Volatile removed in vacuo. Purification of the residue by preparative thin layer chromatography (PTLC) gave the final product.

until the stakes were

Examples 12 to 19

In analogy to the procedure described in Example 9, the following compounds could be prepared:

•O

O'

in Example 1 it was possible

Examples 20 to 22

In analogy to the procedure described, prepare the following compounds:

Pv '4.oco- é> 32_

Claims (13)

Compounds of formula I (I) wherein Z is a tetrapyran group selected from (f) and salts and solvates (for example hydrates) or metabolically labile derivatives of these compounds, wherein: R ^a is C (O) CH ₃ or methyl; R ¹ is hydrogen, halogen, hydroxy, C 1 -C 4 alkoxy or acyloxy; R ² and R ³ are each independently hydrogen, C 1 -C 6 alkyl or C 1 -C 4 alkyl substituted with C 1 -C 4 alkoxy, or R ² and R ³ together with the carbon atom to which they are attached are a carbonyl group (C = O), a thiocarbonyl group (C = S) or a cycloalkyl group having 3 to 8 carbon atoms; R ⁴ is hydrogen or CH ₂ R ⁷ (wherein R ⁷ is hydrogen, hydroxyl, C 1 -C 4 alkoxy or OCOR ⁸ wherein R ⁸ is C 1 -C 4 alkyl or aryl ); R ⁵ and R ⁶ are each independently hydrogen, C 1 -C 6 alkyl or C 1 -C 4 alkoxy substituted with C 1 -C 4 alkyl, or R ⁵ and R ⁶ together with the carbon atom to which they are attached are a carbonyl group (C = O), a thiocarbonyl group (C = S) or a cycloalkyl group having 3 to 8 carbon atoms; n is zero or 1; X and Y are each independently O, S or CR ⁹ R ¹⁰ (wherein R ⁹ and R ¹⁰ are each independently hydrogen, C 1 -C 6 alkyl, C 1 -C 4 alkoxy, or C 1 -C 4 alkyl C 1 -C 4 substituted by C 1 -C 4 alkoxy, or R ⁹ and R ¹¹ together with the carbon atom to which they are attached are a carbonyl group (C = O), a thiocarbonyl group (C = S), a cycloalkyl group containing 3 up to 8 carbon atoms or a group (C 1 CHR ¹¹ where R ¹¹ is hydrogen or C 1 -C 4 alkyl); or when X or Y is an oxygen atom and ⁿ = 0, the groups -Y-CR-R ³ and -X-CR ² R ³ may also be -N = CR ³ - or -NR ¹² -CR ² R ³ - ( wherein CR ² and R ³ are carbonyl (C = O) and R ¹² is C 1 -C 4 alkyl, COR ¹³ where R ¹³ is C 1 -C 6 alkyl) or when Y is oxygen and ⁿ = 0, X may be a CR ¹¹ group (wherein R ¹¹ is as defined above), which is bound by a double bond to the pyran ring; R ¹⁵ is hydrogen, halogen, azido, (C 1 -C 6) alkyl, hydroxy, (C 1 -C 6) alkoxy (optionally substituted with 1 or 2 hydroxy, or ketal thereof, or 1 or 2 C 1 -C 3 alkoxy groups) C 1 -C 4 aryl, C 1 -C 4 aryl-substituted alkoxy, C 3 -C 6 alkenyloxy, OCOR ¹⁸ (wherein R ¹⁸ is C 1 -C 4 aryl-substituted alkoxy or C 1 -C 10 alkyl optionally containing C 1 -C 4 alkyl) one or two double bonds) or C 1 -C 6 alkoxycarbonyl substituted C 1 -C 4 substituted alkoxy and R ¹⁶ is hydrogen or R ¹⁵ and R ¹⁶ together with the carbon atom to which they are attached are a carbonyl group (C = 0) or C = CH _2; R ¹ is CH ₂ R ¹⁹ , wherein R ¹⁹ is hydrogen, hydroxyl, C 1 -C 14 alkoxy, or OCOR ²⁰ , wherein R ²⁰ is C 1 -C 4 alkyl; and W is oxygen, sulfur or CH _2; and the dotted line in group (a) indicates the optional presence of an additional bond; R ^1a is a hydrogen atom, a halogen atom, a hydroxyl group or an alkoxy group having 1 to 4 carbon atoms; R ^2a is a hydrogen atom, a halogen atom, a hydroxyl group or an alkoxy group having 1 to 10 carbon atoms, an alkylthio group having 1 to 10 carbon atoms, an alkoxy group having 1 to 6 carbon atoms, a substituted alkoxy group having 1 to 4 carbon atoms (C 1 -C 6) aryl, (C 3 -C 6) aryl-substituted alkenyloxy, azido, NR ^5a, COR ^5a (wherein each R ^5a is independently hydrogen or (C 1 -C 6) alkyl), OR ^5a (wherein R ^a is a cyclic ether containing 4 to 8 atoms linked to the oxygen atom of the carbon of the cyclic ether which is adjacent to an oxygen atom) or a group Y ^a C (= 0) -X ^a -R ^7a, wherein Y ^is oxygen, a sulfur atom or a group NH, X ^a is either a bond, an oxygen atom or a NR ⁸ group in which R ^9a is a hydrogen atom or a C 1 -C 8 alkyl and R ^7a is C 1 -C 10 alkyl optionally containing one or two double bonds, aryl, aryl substituted C 1 -C 4 alkyl, aryl substituted alkenyl C 2 -C 4 alkyl; C 1 -C 4 haloalkyl, C 1 -C 6 haloalkyl or C 1 -C 6 alkoxy substituted C 1 -C 4 alkyl and R ^3a is hydrogen or R ^2a and R ^3a together with the carbon atom to which they are attached are a carbonyl group (C = O) or a group C = NOR ^9a (wherein R ^9d is an alkyl group having 1 to 6 carbon atoms); and R ^4a is a hydroxyl group, an alkoxy group having 1 to 6 carbon atoms, or an OC (= O) R ^{7a group} (wherein R ^7a is as defined above). A compound according to claim 1 of the structural formula: CN OH A compound according to claim 1 having the structural formula: OH CN HIM OH A compound according to claim 1 of the structural formula A compound according to claim 1 of the structural formula A compound according to claim 1 of the structural formula A compound according to claim 1 of the structural formula: A compound according to claim 1 of the structural formula: 9. A pharmaceutical composition wherein the compound of claim 1 and a pharmaceutically acceptable carrier. 10. An agrochemical composition wherein the compound of claim 1 and an agriculturally acceptable carrier. 11. A method of treating or preventing fungal infections comprising administering a fungicidally amount of a compound of claim 1 to said animals. includes aspects comprising aspects of animals effective 12. A method of controlling phytopathogenic fungi comprising applying a fungicidally effective amount of a compound according to claim 1 to plants in need of such control. 13. A process for producing 4-cyano-4-deformyl-4'-demethylsordarin comprising contacting 4-cyano-4-formylsordarin with a culture of Streptomyces avermitilis in a fermentation medium containing assimilable carbon and nitrogen sources; and isolating 4-cyano-4-deformyl-4 'demethylsordarin from said fermentation medium.